| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
-Subunit Mutations and the Role of Genomic Imprinting
Metabolic Diseases Branch (L.S.W., S.Y., J.L.), National Institute of Diabetes, Digestive, and Kidney Diseases, National Institutes of Health, Bethesda, Maryland 20892; and Department of Molecular, Cellular, and Craniofacial Biology (D.R.W.), University of Louisville School of Dentistry, Louisville, Kentucky 40202
Correspondence: Address all correspondence and requests for reprints to: Dr. Lee S. Weinstein, Metabolic Diseases Branch, National Institute of Diabetes, Digestive, and Kidney Diseases, National Institutes of Health, Bethesda Maryland 20892-1752. E-mail: leew{at}amb.niddk.nih.gov
 |
Abstract |
|---|
-subunit
(Gs) while an intrinsic GTPase activity of
Gs that hydrolyzes bound GTP to GDP leads to
deactivation. Mutations of specific Gs residues
(Arg201 or Gln227) that are critical for the
GTPase reaction lead to constitutive activation of
Gs-coupled signaling pathways, and such somatic mutations
are found in endocrine tumors, fibrous dysplasia of bone, and the
McCune-Albright syndrome. Conversely, heterozygous loss-of-function
mutations may lead to Albright hereditary osteodystrophy (AHO), a
disease characterized by short stature, obesity, brachydactyly, sc
ossifications, and mental deficits. Similar mutations are also
associated with progressive osseous heteroplasia. Interestingly,
paternal transmission of GNAS1 mutations leads to the
AHO phenotype alone (pseudopseudohypoparathyroidism), while maternal
transmission leads to AHO plus resistance to several hormones
(e.g., PTH, TSH) that activate Gs in their
target tissues (pseudohypoparathyroidism type IA). Studies in
Gs knockout mice demonstrate that Gs is
imprinted in a tissue-specific manner, being expressed primarily from
the maternal allele in some tissues (e.g., renal
proximal tubule, the major site of renal PTH action), while being
biallelically expressed in most other tissues. Disrupting mutations in
the maternal allele lead to loss of Gs expression in
proximal tubules and therefore loss of PTH action in the kidney, while
mutations in the paternal allele have little effect on
Gs expression or PTH action. Gs has
recently been shown to be also imprinted in human pituitary glands. The
Gs gene GNAS1 (as well as its murine
ortholog Gnas) has at least four alternative promoters
and first exons, leading to the production of alternative gene products
including Gs, XLs (a novel Gs isoform
that is expressed only from the paternal allele), and NESP55 (a
chromogranin-like protein that is expressed only from the maternal
allele). A fourth alternative promoter and first exon (exon 1A) located
approximately 2.5 kb upstream of the Gs promoter is
normally methylated on the maternal allele and transcriptionally active
on the paternal allele. In patients with isolated renal resistance to
PTH (pseudohypoparathyroidism type IB), the exon 1A promoter region has
a paternal-specific imprinting pattern on both alleles (unmethylated,
transcriptionally active), suggesting that this region is critical for
the tissue-specific imprinting of Gs. The
GNAS1 imprinting defect in pseudohypoparathyroidism type
IB is predicted to decrease Gs expression in renal
proximal tubules. Studies in Gs knockout mice also
demonstrate that this gene is critical in the regulation of lipid and
glucose metabolism. I. Introduction
II. Gs-Signaling Mechanisms and Gene Structure
A. Gs structure and function
B. Gs gene (GNAS1) structure
C. Alternative GNAS1 gene products
D. Imprinting of the GNAS1 gene
III. Gs Activating Mutations
A. Endocrine tumors
B. McCune-Albright syndrome (MAS)
C. Fibrous dysplasia of bone (FD)
IV. Gs Loss-of-Function Mutations
A. Albright hereditary osteodystrophy (AHO)
B. Pseudohypoparathyroidism type IA (PHPIA)
C. Pseudopseudohypoparathyroidism (PPHP)
D. The role of inactivating Gs mutations and
Gs imprinting in the pathogenesis of AHO,
PHPIA, and PPHP
E. Progressive osseous heteroplasia (POH)
V. Insights from Gnas Knockout Mice
A. Tissue-specific imprinting of Gs
B. Role of Gs in renal function
C. Role of Gs in energy and glucose
metabolism
VI. GNAS1 Imprinting Mechanisms and Defects
A. Pseudohypoparathyroidism type IB (PHPIB)
B. Potential GNAS1 imprinting mechanisms
VII. Summary
|
I. Introduction |
|---|
-subunit, which binds guanine
nucleotide and couples to specific receptors and effectors, associated
with a ß- and 
-subunit. The role of G proteins in signal
transduction was last summarized in Endocrine Reviews in
1992 (4).
The first example of a Gs defect leading to
clinical disease was the observation that the secretory diarrhea of
Vibrio cholerae infection results from posttranslational
modification of the Gs -subunit
(Gs) and abnormal regulation of cAMP in
intestinal cells. As Gs is ubiquitously
expressed and is a necessary component for many signaling pathways,
genetic defects of the Gs gene
(GNAS1) would be expected to lead to pleiotropic
manifestations. Within the past decade it has been discovered that
mutations which produce constitutively activated forms of
Gs can lead to endocrine tumors, excess
hormone secretion, and skeletal and other nonendocrine abnormalities.
In contrast, heterozygous inactivating Gs
mutations produce Albright hereditary osteodystrophy (AHO), a syndrome
characterized by skeletal and other developmental abnormalities.
Curiously, maternal inheritance of these inactivating mutations also
produces hormone resistance while paternal transmission does not, and
this is likely due to the fact that Gs is
imprinted (expressed only from the maternal allele) in specific
tissues. Recent studies show that, in fact, GNAS1 is an
extremely complex imprinted gene that produces different gene products
from the maternal and paternal allele through the use of oppositely
imprinted alternative promoters. Moreover, abnormal imprinting of this
gene can also lead to hormone resistance. The diseases associated with
Gs defects are listed in Table 1
.
|
knockout model. Because this article focuses primarily on pathogenesis
rather than on clinical diagnosis and management, we cite references
that describe the clinical aspects of these disorders in greater
detail. |
II. Gs-Signaling Mechanisms and Gene Structure
|
|---|
structure and function-subunit (of which 20 have been identified to date), which
binds guanine nucleotide and interacts with specific receptors and
effectors. ß- And -subunits form tightly but noncovalently bound
dimers that are targeted to the plasma membrane through lipid
modifications at the carboxyl terminus of -subunits. Gs also
undergo lipid modifications that are important for membrane targeting
(9). Association with ß is required for Gs to be
activated by receptors.
Gs is a protein found in all cell types except
mature spermatozoa (10), which couples a wide variety of G
protein-coupled receptors to the stimulation of adenylyl cyclase, the
enzyme that catalyzes the synthesis of cAMP from ATP within cells. Many
of the downstream effects of cAMP are through activation of PKA.
Seven-transmembrane receptors for a wide variety of extracellular
signals, including the glycoprotein and many peptide hormones,
catecholamines, and other biogenic amines and neurotransmitters, all
activate Gs to raise intracellular cAMP in
their target cells. However, Gs activators may
not be limited to the seven-transmembrane receptor family, as there is
evidence that Gs can also be activated by
various tyrosine kinase receptors, including the epidermal growth
factor (11, 12) and basic fibroblast growth factor
receptors (13). Epidermal growth factor receptors appear
to activate Gs by phosphorylation of specific
tyrosine residues in the -subunit (14, 15). Nor is
adenylyl cyclase the only effector activated by
Gs. Gs has been shown
to open specific Ca2+ channels in the heart
(16, 17) and has recently been shown (along with
Gi1) to interact directly with and activate
members of the src tyrosine kinase family (18).
Gs undergoes palmitoylation at its amino
terminus, which is required for membrane targeting
(19, 20, 21). While most attention has been directed at the
role of Gs as a signal transducer at the
plasma membrane, Gs is also present in
intracellular membrane compartments and has been implicated as a
regulator of intracellular membrane trafficking (22). A
second G protein, Golf, also stimulates adenylyl
cyclase, but it is produced by a separate gene and is only expressed in
a small number of tissues (23, 24).
Similar to other G proteins, Gs is activated and
deactivated via the GTPase cycle of its -subunit (Fig. 1) (reviewed in Refs. 4 and
25). In the basal state Gs exists as
an inactive Gs-ß heterotrimer with GDP
bound to the guanine nucleotide binding site of
Gs. Upon interaction with agonist-bound
(activated) receptor, GDP is released from Gs
and replaced with GTP. Binding of GTP to Gs
switches Gs into an active conformation, and
activated GTP-bound Gs dissociates from
ß. Upon activation, Gs is depalmitoylated
and there is evidence that some or all of the -subunit may be
released from the plasma membrane (26, 27, 28, 29). GTP-bound
Gs directly interacts with and activates its
effectors. An intrinsic GTPase activity within the -subunit
hydrolyzes bound GTP to GDP, leading to reassociation with ß to
reform the inactive heterotrimer. While the GTPase activity of other G
protein -subunits is enhanced by interaction with RGS (regulator of
G protein signaling) proteins, there is no evidence for the existence
of an RGS protein for Gs (30, 31). However, there is evidence to suggest that at least one
isoform of adenylyl cyclase (AC5) may enhance the GTPase activity of
Gs and its ability to be activated by
receptors (32). In experimental systems G protein
-subunits can also be activated by incubation with nonhydrolyzable
GTP analogs (e.g., GTPS) or aluminum fluoride
(AlF4-), which binds to
GDP-bound -subunits and mimics the -phosphate of GTP.
|
(33, 34), like all G protein -subunits
(35, 36, 37, 38, 39, 40, 41), have two domains, a ras-like GTPase
domain, which includes the sites for guanine nucleotide binding and
effector interaction, and a more variable helical domain (Fig. 2). Various Gs
isoforms with helical domains of slightly differing length are produced
by alternative exon splicing (42, 43). The guanine
nucleotide resides in a cleft between the two domains, and the helical
domain may be important in preventing the release of GDP from the
-subunit in the inactive state (40, 44). Comparison of
the structures of inactive (GDP-bound) and activated (GTPS- or
AlF4--bound) -subunits
reveals three regions within the GTPase domain (named switch 1, 2, and
3) that undergo conformational changes upon activation. The movement of
switches 1 and 2 upon GTP binding is in direct response to the presence
of the -phosphate group, while switch 3 has no direct contact with
bound guanine nucleotide. Upon activation, switches 2 and 3 move toward
each other and form multiple interactions between acidic and basic
amino acid residues that stabilize the active state
(45, 46, 47, 48). Residues in switch 3 also make contacts with
residues in the helical domain, and these interactions have been
implicated in the maintenance of guanine nucleotide binding (40, 44) and in receptor-mediated activation (44, 49, 50, 51). Two residues in the GTPase domain that are conserved in
all Gs (Arg201 and
Gln227 in Gs, see Fig. 2) are critical for catalyzing the hydrolysis of bound GTP, and
modification or mutation of these residues produces constitutively
active G proteins (37, 38, 52, 53). Crystal structures of
G protein heterotrimers indicate that the G amino terminus and
switch 2 regions are important sites for interaction with the
ß-subunit in the GDP-bound state (33, 39, 41).
|
carboxyl-terminal residues (54, 55, 56, 57, 58), covalent modification
of a specific carboxyl-terminal residue of
Gi/o by pertussis toxin (59),
antibodies directed to G carboxyl termini (60, 61, 62), and
G carboxyl-terminal peptides (63) all prevent or alter
specificity of G protein-receptor coupling. Residues in the 3/ß5
loop near the carboxyl terminus may also directly interact with
receptors and be required for receptor-mediated activation
(58). Mutagenesis studies (64, 65), as well
as crystal structures (34), have also defined the residues
within the GTPase domain of Gs that directly
interact with adenylyl cyclase.
B. Gs gene (GNAS1) structure
The single copy human Gs gene
(GNAS1) is located at 20q13.2–13.3 (66, 67, 68)
while its mouse ortholog (Gnas) is located in a distal
portion of chromosome 2 that is syntenic with human 20q13 (69, 70). The overall structure and regulation of this gene are well
conserved between human and mouse. Originally these genes were defined
by the 13 exons that encode Gs
(43) (Fig. 3). Two long
(Gs-1 and -2) and two short
(Gs-3 and -4) forms of
Gs result from alternative splicing of exon 3,
an exon that encodes a stretch of 15 amino acids within the helical
domain that are not present in other G subunits (42, 43). Use of an alternative splice acceptor site for exon 4 leads
to insertion of an extra serine residue in
Gs-2 and -4. While the ratio of long to short
forms of Gs may vary from tissue to tissue,
there is little evidence to suggest that these alternative
Gs isoforms have distinct roles in signaling
(71), although there may be subtle differences in their
biochemical properties (72). Within the intron between
exons 3 and 4 is an alternative terminal exon (3N, Fig. 3) with its own
polyadenylation signal. Splicing to exon 3N produces transcripts that
are expressed primarily in neural tissues which do not encode
full-length Gs (73). The
importance of these transcripts, if any, is unknown.
|
exon
1 are highly GC rich (43, 74) and are contained within a
CpG island, which is typical for promoters of ubiquitously expressed
genes such as Gs [CpG islands are 
1- to
3,000-bp regions that are highly GC rich with a relatively high
frequency of CpG dinucleotides and are usually unmethylated (75, 76)].
The GC-rich Gs promoter region is unmethylated
in both human and mouse (74, 77, 78) (see Section
II.D).
It is now known that both GNAS1 and Gnas are more
complex genes containing at least four alternative promoters and first
exons that splice onto a common exon (exon 2). The most downstream of
these exons is Gs exon 1 (Fig. 3
). The most
upstream alternative promoter (located 49 kb upstream of
Gs exon 1) produces transcripts encoding the
chromogranin-like protein NESP55 (78, 79, 80). The entire
coding sequence for NESP55 is contained within the upstream exon, and
therefore Gs exons 2–13 are within the
3'untranslated region (3'-UTR) of the NESP55 transcripts
(79, 81). There is some evidence in humans for the
presence of a minor splice donor site within the 5'-UTR of the NESP55
exon. In the orthologous exon in mouse (referred to as
Nesp), the presence of both splice donor and acceptor sites
leads to the presence of a 95-bp intron, which disrupts the 5'-UTR of
Nesp. In both human and mouse, the NESP55 promoter region is
methylated only on the paternal allele, and NESP55 is only transcribed
from the maternal allele (78, 79, 82) (see Section
II.D).
A third alternative promoter located approximately 11 kb downstream of
the NESP55 exon and about 35 kb upstream of Gs
exon 1 in humans produces transcripts encoding XLs, an isoform of
Gs with a long amino-terminal extension
(77, 78, 80, 83) (Fig. 3). The alternative amino-terminal
region of XLs is encoded by its first exon, while the
carboxyl-terminal portion of the protein, which is identical to
Gs, is encoded by exons 2–13. Just downstream
of the XLs exon are two small exons of 91 and 67 bp in length
(referred to as A20 and A21, Ref. 77). While most XLs
transcripts do not contain exons A20 and A21, a small proportion of
XLs transcripts have A20 alone (84) or both A20 and A21
(77) spliced in between the upstream XLs exon and exon
2. The presence of these exons within the spliced mRNA disrupts the
coding sequence of XLs. mRNAs that contain the A20 sequence produce
a carboxyl-terminally truncated form of XLs called XLN1b (see Fig. 3
and Ref. 84). A role for these transcripts, if any,
remains to be determined. It is not known whether exons comparable to
A20 and A21 exist in the orthologous region in mouse (which is referred
to as Gnasxl).
The XLs promoter region is located within the 3'-half of a large
6-kb CpG island. In both humans and mice, XLs is only transcribed
from the paternal allele and its promoter is methylated only on the
maternal allele (77, 78, 82). The region between the
NESP55 and XLs promoters is well conserved between human and mice
(85). In particular, a highly conserved region located
approximately 2–3 kb upstream of the XLs exon is a promoter for
antisense transcripts that traverse the NESP55 exon from the opposite
direction (82, 85) (Fig. 3). In humans, a large variety of
antisense mRNA transcripts result from alternative splicing of five
exons (one upstream and four downstream of the NESP55 upstream exon)
and the use of multiple splice sites (85). In the mouse,
the transcripts (which have been named Nespas) originate
from the same location, but the exact exon structure has not been well
defined (82, 85, 86). Unlike the case in humans, the mouse
Nespas mRNA product incorporates the complementary sequence
spanning the upstream Nesp exon (86). Similar
to XLs, the NESP55 antisense transcripts are only expressed from the
paternal allele, and their promoter is methylated on the maternal
allele (82, 85, 86).
A fourth promoter that generates transcripts from the sense strand is
located approximately 2.5 kb upstream of Gs
exon 1 (Fig. 3). The alternative first exon [which has been called A/B
in humans (87) and 1' in dogs (88), and which
we call 1A (74, 89)] splices onto exon 2 through the use
of two alternative splice donor sites. There is no consensus AUG
translational start site within exon 1A, and the resulting exon 1A
transcripts are presumed to be untranslated mRNAs (74, 88). These transcripts are ubiquitously expressed in mouse, and
have a tissue distribution pattern that is very similar to that of
Gs (74). The exon 1A promoter
region is located within a CpG island that appears to be distinct from
the Gs exon 1 CpG island (74, 89). In both humans and mice, the exon 1A promoter is normally
only methylated on the maternal allele, and exon 1A mRNAs are only
transcribed from the paternal allele (74, 89). As
discussed below, imprinting of this region is lost in patients with
pseudohypoparathyroidism type IB (PHPIB), and this region is probably
important for the tissue-specific imprinting of
Gs.
C. Alternative GNAS1 gene products
1. XLs. XLs is an isoform of Gs
in which the first 47 amino acids encoded by
Gs exon 1 are replaced by a large
amino-terminal region encoded by XLs exon 1 (the XL domain) to
produce an acidic 78-kDa protein that has an electrophoretic mobility
of 94 kDa (83, 84) (Fig. 3). Within the XL domain are, in
sequential order, a region of Glu-Pro-Ala-Ala (EPAA) repeats, a region
of Ala-Ala-Arg-Ala (AARA) repeats, a proline-rich region, and a
cysteine-rich region (83, 84). The carboxyl-terminal
portion of the XL domain is similar to that of the carboxyl-terminal
portion of Gs exon 1, a region required for
guanine nucleotide binding and ß association. There is also
evidence for the in vivo expression of carboxyl-terminally
truncated forms of XLs resulting from the splicing of exon 3N to
exon 3 (XLN1a) or the insertion of exon A20 (XLN1b) or exons A20 and
A21 (see Refs. 77 and 84 and Fig. 3). The
biological significance of these so-called XLN1 isoforms is
unknown.
While RT-PCR experiments suggest that XLs is widely distributed
(82), Northern analysis, immunoblotting, and in
situ hybridization experiments demonstrate that XLs expression
is limited to neural and endocrine tissues (83, 84).
XLs is expressed at very high levels in rat pituitary (particularly
in the melanotrophs within the pars intermedia of the posterior lobe),
and to a lesser extent in brain, adrenal, heart, and pancreatic islets
(where it is expressed in only some cells and is not restricted to
ß-cells). There is little or no XLs expressed in spleen, liver, or
kidney. In the rat, expression of XLs begins during the onset of
neurogenesis (90). While initial sucrose gradient studies
suggested that XLs is primarily localized to the trans-Golgi network
(83), more recent sucrose gradient and immunofluorescence
studies demonstrate that XLs is mostly targeted to the plasma
membrane (84). However, one study suggests that XLs (at
least when transfected into cells) may also be targeted to Golgi under
certain conditions (91). It is clear that the XL domain is
critical for membrane targeting (84, 91). Palmitoylation
of cysteines within the cysteine-rich region is required for membrane
association, and the proline-rich region is required for targeting to
Golgi (91).
Biochemical studies indicate that XLs has many properties in common
with Gs (92). Like
Gs, XLs can be modified by cholera toxin,
can interact with ß, and is activated by the nonhydrolyzable GTP
analog GTPS. Also similar to Gs, XLs
proteins that are activated by GTPS or by mutating residue
Gln548 (analogous to residue
Gln227 in Gs which when
mutated leads to reduced GTPase activity and constitutive activation)
can stimulate adenylyl cyclase activity. However, in contrast to
Gs, there is no evidence that XLs can be
activated by seven-transmembrane receptors that activate
Gs (92). Therefore, perhaps
XLs is a stimulator of adenylyl cyclase that is activated by an
alternative pathway. The biological function of XLs remains to be
determined.
2. NESP55. NESP55 (neuroendocrine-specific protein of apparent molecular mass 55 kDa) is an acidic chromogranin that localizes to large dense-core granules within neuroendocrine tissues, such as the adrenal medulla, anterior and posterior pituitary, hypothalamus, and other brain regions (81, 93). Within the adrenal medulla, NESP55 is more highly expressed in epinephrine-, as opposed to norepinephrine-secreting chromaffin cells (94). Within the central nervous system, NESP55 is expressed in neural, but not glial, cells and is most prominent in the pituitary, hypothalamus, and other midbrain and brainstem regions, but absent in the neocortex, hippocampus, and cerebellum (95). Its brain distribution overlaps the distribution pattern of the norepinephrine, epinephrine, and serotonergic transmitter systems. There is also some NESP55 expression in the intestine, where it undergoes extensive proteolytic cleavage (93). The NESP55 protein is highly conserved between species and is a predicted size of approximately 27–29 kDa (96). However, it is very acidic due to addition of keratin sulfate glycosaminoglycan chains (and therefore has an apparent molecular mass of 55 kDa on immunoblots) (81, 96) and undergoes proteolytic cleavage. During midgestation in the mouse, Nesp transcripts are expressed in somites and vasculature (97). Little is known about the physiological role of NESP55, although it has been proposed that one potential proteolytic cleavage product may be an endogenous antagonist of serotonin 1b receptors (81). Loss of NESP55 expression in humans is not associated with an obvious phenotype (89).
D. Imprinting of the GNAS1 gene
Genomic imprinting is an epigenetic phenomenon affecting a small
number of autosomal genes (probably 100 or less) that leads to
differences in gene expression between the maternal and paternal allele
(98, 99, 100, 101). Mutation or dysregulation of imprinted
genes is associated with several human congenital diseases, including
Beckwith-Wiedemann, Prader-Willi, and Angelman’s syndromes, and in
carcinogenesis. The first clue to the existence of imprinting was the
observation that both androgenetic and parthenogenetic zygotes develop
abnormally, demonstrating that normal development requires both a
paternal and maternal contribution to the genome (102, 103). Subsequently, specific chromosomal regions in the mouse
were identified that were presumed to contain imprinted genes due to
the fact that uniparental disomy (UPD) of these regions produced
abnormal phenotypes (104, 105). (UPD is the inheritance of
a chromosome or chromosomal region from a single parent.) Often these
imprinted regions contain several closely linked imprinted genes that
are coordinately regulated. One such imprinted region is in the distal
portion of chromosome 2 and includes Gnas (70, 104).
In order for the parental alleles of imprinted genes to behave differently, there must be a mark placed on the gene in either the male or female gamete that is maintained in all somatic cells throughout development. This mark would have to be erased in the primordial germ cell so that the appropriate imprinting patterns could be reestablished in the mature oocyte or spermatozoa, respectively. Virtually all imprinted genes have one or more regions in which the cytosines within CpG dinucleotides are methylated on only one parental allele, and differential methylation is the most likely candidate for the imprinting mark (99, 101). Imprinting is lost in mice lacking the DNA methyltransferase DNMT1 (106, 107). In many instances, methylation within the differentially methylated regions (DMRs) is established in either the male or female gamete and maintained throughout pre- and postimplantation development, and deletion of these DMRs disrupts the imprinting of one or more genes in the imprinted region (108, 109, 110, 111, 112). As differential methylation of these regions is probably important for establishing imprinting, these regions have been referred to as methylation imprint marks or core DMRs. In contrast, other regions become differentially methylated at a later time in development, and while these regions may be important to maintain imprinted expression, they do not provide the initial signal that marks parental origin.
In many, but not all, cases, DMRs coincide with the gene promoter and lead to allele-specific expression through silencing of the methylated promoter (98, 99, 113). This occurs through the binding of methyl-CpG binding proteins, which recruit histone deacetylases, leading to a more compact chromatin that is not accessible to transcriptional factors (99, 101, 113, 114). There is also recent evidence that the DNA methyltransferases also recruit histone deacetylases (115, 116). Methylation itself may prevent the binding of transcriptional factors that are required for gene transcription. However, differential methylation of imprinted genes can lead to allele-specific expression through alternative mechanisms. For example, the H19 DMR is located between the Igf2 promoter and a set of endoderm-specific enhancers and contains several boundary elements (or insulators). On the maternal allele the DMR is unmethylated and is therefore able to bind the insulator protein CCCTC-binding factor (CTCF) and insulate the Igf2 promoter from the enhancers. As a result, Igf2 is not expressed from the maternal allele. On the paternal allele the DMR (and its boundary elements) are methylated. CTCF is unable to bind to the methylated boundary elements. Consequently, the DMR does not insulate the Igf2 promoter from its enhancers, allowing Igf2 to be expressed from the paternal allele (117, 118). A similar mechanism has been recently proposed for the DLK1-GTL2 imprinting cluster (119, 120).
GNAS1 has a complicated imprinting pattern that leads to the
expression of some gene products from the maternal allele and others
from the paternal allele (Fig. 3). The imprinting pattern of the mouse
ortholog Gnas is essentially identical to that of
GNAS1. The closely linked promoter regions for NESP55 and
XLs are oppositely imprinted. NESP55 and its associated transcripts
are only expressed from the maternal allele, and its promoter region is
methylated on the paternal allele (78, 79, 82). In
contrast, XLs and its associated transcripts are only expressed from
the paternal allele, and its promoter region is methylated on the
maternal allele (77, 78, 82). Based on the similar tissue
distribution but opposite imprinting patterns, it is possible that
these two imprinted regions are coordinately regulated.
Paternal-specific methylation of the NESP55 promoter is not established
until postimplantation development in mouse, and therefore this region
is not the methylation imprint mark for the Gnas locus
(74). More likely its imprinting is dependent on another
imprinting event that occurs at an earlier stage in development.
The NESP antisense transcript is expressed only from the paternal
allele, and its promoter (which is located in the same CG-rich region
as the XLs promoter) is methylated on the maternal allele (82, 85, 86). Interestingly, paternal-specific antisense transcripts
have been identified in several imprinted genes (111, 121, 122, 123). One potential mechanism for the imprinting of NESP55
is that the paternal antisense transcript leads to silencing of the
NESP55 promoter on the paternal allele. This is exactly analogous to
the situation in the maternally expressed Igf2r gene
(111). Downstream of the Igf2r promoter is a
promoter for a paternal-specific antisense transcript that is
methylated on the maternal allele. The maternal-specific methylation of
the antisense promoter is established in the oocyte and is maintained
throughout development and is therefore a methylation imprint mark,
whereas methylation and silencing of the paternal Igf2r
promoter is established later in development. Moreover, deletion of the
differentially methylated antisense promoter region leads to loss of
imprinting of Igf2r. Further studies will be required to
determine when methylation of the NESP antisense promoter is
established and whether or not it is required for NESP55 imprinting.
One study suggested the NESP antisense expression is biallelically
expressed in two tissues (82). However, one of these
tissues was testis, where the transcript would be predicted to be
biallelically expressed because the promoter is unmethylated in male
germ cells. An in situ hybridization study of midgestational
mouse embryos showed that Nesp and Nespas
transcripts do not colocalize, indicating that at least during
gestation Nespas probably does not play a role in the
imprinting of Nesp (97).
The temporal methylation pattern of the XLs promoter region in mice
is atypical as the region appears to be partially methylated in both
spermatozoa and oocytes, and this methylation is maintained in
preimplantation blastocysts (at a time when the genome is undergoing
global demethylation) (74). These findings suggest that
allele-specific methylation is not erased in germ cells and that
paternal-specific methylation is somehow reestablished at some point
during postimplantation development by a novel mechanism. The role of
the XLs promoter region in establishing GNAS1 imprinting
is presently undefined.
Clinical genetic studies of AHO patients (8, 124) strongly
suggest that Gs is imprinted in a
tissue-specific manner, being biallelically expressed in most tissues
but maternally expressed in a few tissues, such as the renal proximal
tubules, and this has been confirmed in Gnas knockout mice
(125). Although earlier studies did not show imprinting of
Gs in humans (77, 79, 126),
Gs has recently been shown to be imprinted in
normal human pituitary glands (127). Even in tissues where
Gs is imprinted, the imprinting is not
absolute in that there is some expression from the paternal allele
(125, 127). The Gs promoter is
not methylated in either allele, and therefore allele-specific
expression of Gs is not due to differential
methylation of its promoter (74, 77, 78). NESP55 and
XLs are expressed in tissues where Gs is
not imprinted (77, 79), while neither is expressed in
tissues where Gs is imprinted (Refs. 84, 93 and 128 and S. Yu and L. S. Weinstein,
unpublished data). It is therefore unlikely that either NESP55 or
XLs or their promoters are involved in the imprinting of
Gs. Tissue-specific imprinting of
Gs and the role of Gs
imprinting in human disease are discussed in more detail in
Sections IV.D, V.A, and VI.
The exon 1A promoter region is methylated on the maternal allele, and
exon 1A mRNAs are only expressed from the paternal allele (74, 89). In mice the maternal-specific methylation of the exon 1A
promoter is established in the oocyte and maintained throughout pre-
and postimplantation development (74); therefore, this
region appears to be a methylation imprint mark that may be important
to establish imprinting throughout the GNAS1 locus. Like the
NESP antisense transcript, the exon 1A mRNAs are untranslated
mRNAs. Untranslated mRNAs are a common feature of imprinted genes,
although their role in the imprinting mechanism, if any, is unknown.
The close proximity of the exon 1A DMR to the
Gs promoter and the abnormal imprinting of
this region in PHPIB (89), a disease likely to result from
abnormal imprinting of Gs, strongly suggests
that this region is critical for the tissue-specific imprinting of
Gs. Potential mechanisms by which this region
may lead to Gs imprinting are discussed in
Section VI.B.
As imprinted genes often occur in clusters, it is important to identify the genes most closely linked to GNAS1 and determine whether or not they are also imprinted. The two closest genes located downstream of GNAS1 are TH1, a gene of unknown function, and CTSZ, which encodes cathepsin Z, and initial results suggest that neither gene is imprinted (129). The imprinting status of neighboring genes upstream of GNAS1 remains to be established. In the mouse, 14 genes close to Gnas that are located within the 20-centimorgan distal chromosome 2 imprinted region were shown not to be imprinted (130, 131).
|
III. Gs Activating Mutations
|
|---|
activation can lead to pathophysiological consequences was provided by
the fact that the secretory diarrhea characteristic of intestinal
V. cholerae infection results from a covalent modification
of Gs that leads to constitutive activation.
V. cholerae secretes an exotoxin named cholera toxin that
catalyzes the ADP ribosylation of a specific
Gs residue (Arg201) that
is critical for the G protein’s GTPase "turn off" mechanism
(4). As a result of markedly reduced GTPase activity,
Gs remains in its active GTP-bound form for a
long period of time, resulting in constitutive Gs
activation (Fig. 1). Excess intracellular cAMP in intestinal cells
alters ion transport, leading to secretory diarrhea. Subsequently, it
has been established that missense mutations that encode substitutions
of Arg201 or residue Gln227
also lead to decreased GTPase activity and constitutive activation
(52, 53, 132) (Fig. 2), and such mutations have been
identified in sporadic endocrine tumors, the McCune-Albright syndrome
(MAS), and fibrous dysplasia of bone (FD).
A. Endocrine tumors
In many endocrine glands, cAMP stimulates both proliferation and
hormone secretion, and therefore mutations that lead to increased cAMP
levels, such as activating Gs mutations, would
be predicted to lead to differentiated endocrine tumors with increased
rates of hormone release. The first step toward the identification of
activating Gs mutations in a human disease was
the observation that a subset of human pituitary GH-secreting tumors
have high basal levels of GH release and membrane-associated adenylyl
cyclase activity that could not be further stimulated by GHRH, which
normally stimulates adenylyl cyclase through its
Gs-coupled receptor (133). These
tumors, which account for 40% of GH-secreting tumors, were shown to
contain heterozygous missense mutations in GNAS1 exons 8 or
9 that encode substitutions of Gs residues
Arg201 (to Cys or His) or
Gln227 (to Arg or Lys), respectively (52, 134). Subsequently, a tumor containing an
Arg201-to-Ser mutation was identified
(135). These mutations were of somatic origin, as the same
mutations were not present in peripheral blood samples from the same
patients. Mutation of either Arg201 or
Gln227 leads to decreased intrinsic GTPase
activity (52, 53, 132), and the catalytic role of these
amino acid residues in the GTPase reaction is supported by data from
x-ray crystallography (37, 38) (Fig. 2). The mutated
Gs is therefore constitutively activated,
leading to activation of adenylyl cyclase and further downstream
physiological responses. In GH-secreting pituitary cells, cAMP
stimulates proliferation and differentiation by inducing the
tissue-specific transcription factor GHF1 (136).
Consistent with these findings in human GH-secreting tumors, transgenic
mice expressing cholera toxin (which covalently modifies
Gs Arg201) in
their somatotrophs developed pituitary hyperplasia and excess GH
secretion (137). Activating mutations are dominant acting,
and such constitutively activated forms of Gs
have been designated as the gsp oncogene (52, 134).
In gsp+ GH-secreting pituitary tumors, mRNA derived from the
gsp+ allele is often more abundant than mRNA derived from
the wild-type allele (52, 127, 138). A recent study showed
that Gs is imprinted and expressed from the
maternal allele in normal pituitary glands and that in 21 of 22
gsp+ GH-secreting tumors the gsp mutation was
present on the maternal allele (127). Presumably, the
gsp mutation only leads to significant expression of
activated Gs (and therefore tumorigenesis)
when present on the active maternal allele (127, 139). In
both gsp+ and gsp- GH-secreting tumors, there is
variable loss of Gs imprinting leading to a
variable amount of Gs expression from the
normally inactive paternal allele (127). Whether this is
critical for tumorigenesis or represents an epiphenomenon and whether
loss of Gs imprinting in pituitary tumors is
associated with altered GNAS1 methylation remain to be
determined (139).
There are few clinical differences between gsp+ and
gsp- GH-secreting tumors (140, 141).
Gsp+ tumors tend to be smaller than gsp- tumors
and maintain their responsiveness to somatostatin analogs in terms of
inhibition of GH secretion (135, 142, 143). This indicates
that Gi pathways, which inhibit adenylyl cyclase,
are preserved and that somatostatin analogs are useful therapeutic
agents for gsp+ GH-secreting tumors. There are several
mechanisms that counteract the effect of gsp mutations on
intracellular cAMP levels. The levels of Gs
protein are reduced in gsp+ pituitary tumors, presumably due
to more rapid turnover of activated Gs
proteins (138). Also, Gs
activation in gsp+ tumors leads to increased levels of a
specific subtype of phosphodiesterase (phosphodiesterase 4), which
hydrolyzes cAMP and therefore counteracts the increased rate of cAMP
formation (144). Phosphodiesterase 4 expression was also
elevated in one gsp+ thyroid tumor (145).
Gsp mutations are also rarely present in other pituitary tumors. Similar to GHRH in somatotrophs, CRH activates Gs in corticotrophs to stimulate ACTH release, and therefore one might predict gsp mutations to be present in corticotroph adenomas. One study identified gsp mutations in 2 of 32 corticotroph tumors (146), while another study failed to find mutations in 7 such tumors (134). It is interesting to note that corticotroph tumors are not a feature of MAS, in which the distribution of gsp mutations is widespread. Results from three studies indicate that gsp mutations are also present in about 10% of clinically nonfunctional pituitary adenomas (134, 147, 148). Studies have failed to identify gsp mutations in either thyrotroph or lactotroph tumors (134, 149). This is not surprising given the fact that their respective hypothalamic stimulating hormones do not work through Gscoupled pathways.
Constitutively activated Gs transfected into
the FRTL5 thyroid cell line leads to increased growth and thyroid
hormone release (150), and transgenic mice with
thyroid-specific expression of cholera toxin develop thyroid
hyperplasia and hyperthyroidism (151). In thyroid cells,
cAMP stimulates both cAMP-dependent transcription factors and the p38
MAPK in a PKA-dependent manner (152). Several studies have
confirmed that gsp mutations are present in a small subset
of autonomously functioning thyroid tumors (134, 153).
More common are activating TSH receptor mutations, which activate the
same Gs-coupled pathway (154, 155).
The gsp mutations are also rarely found in differentiated
thyroid carcinomas (153, 156, 157). One study showed that
follicular neoplasms have increased levels of presumably normal
Gs (158). While gsp
mutations have been identified in parathyroid and adrenocortical tumors
and in pheochromocytomas (159, 160), other studies suggest
that these mutations are rarely found in these or other endocrine
tumors (134, 160, 161, 162).
B. McCune-Albright syndrome (MAS)
MAS was first described by McCune (163) and Albright
and co-workers (164) in the 1930s and is classically
defined as the concurrence of hyperpigmented (café-au-lait) skin
lesions, sexual precocity, and FD (for reviews, see Refs.
6 and 165, 166, 167, 168, 169). MAS patients may also
present with other endocrine or nonendocrine abnormalities. Some MAS
patients present with only two features of the classic triad or one of
these features and another endocrine or nonendocrine abnormality
(170, 171, 172). In virtually every case, MAS occurs
sporadically and does not appear in the subsequent generation.
The café-au-lait spots in MAS are single or multiple tan-brown hyperpigmented flat macules with irregular ("coast of Maine") borders (164) that become more obvious with age or with sun exposure (172, 173). Often these lesions are limited to one side of the body, which usually corresponds to the side with bone involvement and generally do not cross the midline. They are often arranged in a segmental pattern, which follows the developmental lines of Blaschko (174). Melanocytes cultured from these lesions have increased intracellular cAMP levels, increased numbers of dendrites and melanosomes, and increased levels of tyrosinase, the rate-limiting enzyme for the production of melanin (175). In MAS patients usually more than one bone is affected by FD, which is described in more detail in the next section.
In female MAS patients the first sign of sexual precocity is premature menses, sometimes occurring within the first months of life (172, 173, 176). Serum estrogen levels fluctuate, with peaks corresponding to the presence of large ovarian follicles (171, 176, 177). Usually females undergo normal development during adolescence and show normal reproductive function in adult life. The typical signs of sexual precocity in males include testicular enlargement and premature onset of secondary sex characteristics and spermatogenesis (172, 173). In one case, testicular enlargement in the absence of pubertal development was due to premature maturation of the seminiferous tubules but not of the Leydig cells (178).
Other endocrine abnormalities associated with MAS include the presence of hyperfunctional thyroid nodules, usually leading to hyperthyroidism (172, 173, 179, 180, 181), macronodular adrenal hyperplasia or adrenal adenomas leading to hypercortisolism (172, 173, 182, 183), pituitary tumors leading to acromegaly and/or hyperprolactinemia (172, 173, 184), and hypophosphatemic rickets or osteomalacia (172, 173, 185). In each case the peripheral endocrine organs (thyroid, adrenal cortex, gonads) behave as if they are being overstimulated by the pituitary even though circulating levels of their respective stimulatory pituitary hormone (e.g., TSH, ACTH, gonadotropins) are suppressed (172, 173, 179, 180, 186, 187, 188, 189). Pituitary adenomas and hyperplasia have not been associated with excess secretion of other pituitary hormones in MAS.
Hyperphosphaturic hypophosphatemic rickets or osteomalacia has been associated with MAS and polyostotic fibrous dysplasia (POFD) (172, 173, 190, 191, 192, 193, 194, 195, 196). In one study increased phosphate excretion was documented in almost 50% of patients with MAS or POFD (196). Some have postulated that hyperphosphaturia in MAS is due to a phosphaturic factor (phosphatonin) secreted from areas of fibrous dysplasia, similar to the mechanism of tumor-associated hypophosphatemic rickets (191, 194, 196), while others implicate a primary defect in the renal proximal tubule (193, 195). Increased intracellular cAMP levels in renal proximal tubules, even in the absence of hyperparathyroidism, leading to decreased phosphate reabsorption, could be one potential mechanism for hypophosphatemia in MAS patients. One study showed that MAS patients with hypophosphatemia have increased basal levels of urinary cAMP (197), while another study reported normal levels of urinary cAMP in these patients (196). In addition to increased phosphate clearance, patients with MAS or POFD have other evidence of renal tubular dysfunction, including aminoaciduria and mild proteinuria (196). These abnormalities are similar to those observed in patients with tumor-induced osteomalacia, who presumably develop renal abnormalities due to excess circulating levels of a phosphatonin [possibly fibroblast growth factor 23 (198)]. Although hyperparathyroidism has been reported to occur in patients with monoostotic fibrous dysplasia (MOFD) usually affecting the jaw (199, 200, 201), it is likely that most or all of these patients have hyperparathyroidism-jaw tumor syndrome that has been mapped to chromosome 1 (202, 203). Hyperparathyroidism is rarely present in patients with the classic MAS clinical triad (204, 205).
The abnormalities in MAS patients are generally restricted to bone, skin, and endocrine organs, and therefore there is little effect on mortality (172, 206, 207). However, some patients also develop one or more nonendocrine abnormalities that may markedly increase morbidity and mortality (208). Often these patients also have extensive POFD or hypercortisolism (206, 208). Nonendocrine organs that may be affected in MAS include the liver, heart, thymus, spleen, bone marrow, gastrointestinal tract, and brain (208, 209). Liver abnormalities associated with MAS include severe neonatal jaundice, elevated liver enzymes, and cholestatic and biliary changes on liver biopsy (208, 210). Cardiac abnormalities may include cardiomegaly associated with atypical myocyte hypertrophy, persistent tachycardia, and unexplained sudden death (208). Other rare features of MAS include thymic hyperplasia, myelofibrosis, gastrointestinal polyps, pancreatitis, breast cancer, microcephaly, and other neurological abnormalities (164, 172, 208, 211, 212, 213).
Based on the fact that the cutaneous hyperpigmentation in MAS often follows the developmental lines of Blaschko, Happle proposed that MAS is likely to be caused by a dominant somatic mutation occurring early in development or a gametic half-chromatid mutation (174). Either of these early mutational events would produce a mosaic with a widespread distribution of mutant cells. Because MAS is never inherited, it is likely that the underlying mutation is lethal if present in the germline. According to this model, the specific constellation of abnormalities within a given patient would be dependent on the distribution of mutant-bearing cells.
Although it was originally proposed that the increased growth and
secretory function of diverse peripheral endocrine organs in MAS
results from a hypothalamic defect (214), it is now clear
that each of the peripheral endocrine glands functions autonomously in
the absence of its respective trophic pituitary hormone (173, 179, 186, 187, 188, 189). These trophic hormones (e.g.,
gonadotropins, TSH, ACTH, GHRH) normally stimulate
Gs and cAMP generation in their respective target
tissues, leading to increased proliferation and hormone secretion
(173, 188, 207, 215). Moreover, -MSH normally raises
cAMP levels in melanocytes by binding to the
Gs-coupled melanocortin 1 receptor, leading to
increased tyrosinase activity and melanin production (216, 217). A genetic lesion that leads to increased cAMP generation,
even in the absence of stimulating hormone, such as a gsp
mutation, could explain the endocrine and skin manifestations of MAS.
This idea was supported by biochemical studies that showed cAMP
production to be increased in tissues isolated from MAS patients
(175, 197, 218, 219).
The gsp mutations coding for Gs
Arg201 substitutions have been identified in
tissues from many MAS patients (208, 209, 220, 221). In
each case, one specific mutation (either Arg201
to His or Cys) is detected in multiple tissues in variable abundance,
consistent with a somatic mutation with a widespread distribution. More
recently, an Arg201 to Gly mutation was
identified in one MAS patient (222), and an
Arg201 to Leu mutation, which was possibly
germline, was identified in a patient with severe skeletal, endocrine,
and developmental abnormalities (223). The gsp
mutations coding for Gs
Gln227 substitutions have not been found in MAS,
perhaps because these mutations are more activating than
Arg201 mutations (52) and therefore
may lead to a more lethal phenotype. Consistent with the mutation being
dominant, the wild-type allele is always present in equal or greater
abundance than the mutant allele, indicating that the affected cells
are heterozygous for the gsp mutation.
The gsp mutations have been identified in affected endocrine tissues and in café-au-lait lesions from MAS patients (175, 208, 209, 221), and it is likely that these lesions are the result of increased intracellular cAMP levels. gsp Mutations have also been identified in other normal and abnormal nonendocrine tissues (208, 209, 210). It seems possible that increased activity of Gs pathways, which normally mediate the chronotropic and ionotropic responses to sympathetic stimulation, could lead to the cardiac hypertrophy and sudden death occasionally found in MAS patients (208). Some of these effects on cardiac function may result from increased PKA-dependent activation of p38 MAPK (224). The role of gsp mutations in the pathogenesis of these and other nonendocrine manifestations needs to be defined.
The clinical features present in an individual MAS patient are
primarily determined by the distribution of cells that bear the
gsp mutation. Patients who present with only one or two
features of MAS have the same somatic mutations, but in a more limited
tissue-specific distribution (presumably these mutations occur at a
later time in development). Given that in some tissues
Gs is preferentially expressed from the
maternal allele, MAS patients with gsp mutations in the
maternal allele are perhaps more severely affected than those patients
with mutations in the paternal allele. Support for this comes from the
fact that in gsp+ GH-secreting tumors, the gsp
mutation is almost always on the maternal allele (127).
Activating mutations within the paternally expressed XLs protein
also lead to constitutive activation of adenylyl cyclase
(92). Whether activated XLs contributes to the clinical
manifestations in MAS patients with paternal gsp mutations
is unknown.
C. Fibrous dysplasia of bone (FD)
FD is a focal and benign fibrous bone lesion that was first
described about 50 yr ago (225). Most FD patients
(70%) have a single bone lesion (MOFD), while the remainder have
multiple lesions (POFD) (226). A small minority of POFD
patients have other features of MAS. With rare exceptions
(227, 228, 229), FD is a sporadic disease that is almost never
inherited. FD lesions are typically found in the long bones of the
extremities, the ribs, and craniofacial bones and are rarely found in
the hands, feet, or spine (206, 226, 230). FD lesions may
be asymptomatic but often lead to bone deformity, pain, pathological
fractures, or cranial nerve compression (172, 206, 230).
In some patients FD is associated with intra-muscular myxomas
(Mazabraud’s syndrome) (193, 231, 232, 233), and
gsp mutations have been identified in intramuscular myxomas
both in patients with and without accompanying FD (233).
The clinical, radiological, and histological features of FD are
reviewed in Refs. 165 and 166 .
FD is a lesion composed mainly of fibrous tissue that originates in the medullary cavity and expands concentrically outward into the surrounding cortical bone. Most of the cells are immature mesenchymal cells with a spindle-shaped fibroblastic appearance (234). These cells express alkaline phosphate and other osteoblast-specific proteins (235). Within the fibrous tissue are spicules of immature woven bone. The histological pattern of FD varies depending on location, but all lesions have common characteristic features (236). For example, the woven bone is lined by flat cells (234) with retracted cell bodies, leading to the formation of pseudolacunar spaces (235, 236). This retraction is likely to be the result of increased intracellular cAMP, as cAMP induces retraction in cultured osteoblasts (235). In addition, in FD the collagen fibrils are perpendicular to the bone-forming surface (so-called Sharpey’s fibers) (235, 236, 237). In some FD lesions, islands of hyaline cartilage are present, and this can sometimes be a dominant feature (234, 238). Often FD lesions have osteomalacic changes, even in patients without hypophosphatemia (237).
FD originates in the inner marrow cavity and expands into surrounding cortical bone through the bone-resorbing activity of osteoclasts, which are present in greater numbers in the periphery (218, 235). Bisphosphonates have been shown to have therapeutic benefit, presumably due to their antiresorptive activity (239, 240, 241, 242). These lesions rarely expand beyond the normal boundaries of the cortical bone (243, 244) or undergo malignant degeneration (245, 246, 247, 248).
As in MAS, activating Gs mutations have also
been found in FD lesions from MAS, POFD, and MOFD patients, indicating
that all forms of FD have the same underlying genetic abnormality
(219, 220, 237, 249, 250, 251, 252). In most cases the mutation
encodes Arg201 to Cys or
Arg201 to His substitutions, although an
Arg201 to Ser mutation was found in one POFD
patient (252).
Recent studies suggest that FD results from abnormal proliferation and differentiation of bone marrow stromal cells (253). Under normal conditions, marrow stromal (CFU-F) cells differentiate into mature osteoblasts through a series of defined steps: a proliferative phase in which precursor cells divide and a postproliferative phase in which the cells differentiate and express osteoblast-specific gene products (e.g., alkaline phosphatase, osteopontin, and osteocalcin) that support the formation of mineralized bone (254). Cells isolated from FD lesions have an increased proliferation rate and are poorly differentiated, expressing early, but not late, markers of osteoblast differentiation (219, 235). Transplantation of these cells into immunocompromised mice produces fibrotic lesions that are similar to human FD, with abnormal woven bone ossicles and no interspersed adipocytes or hematopoietic elements (237, 255), further suggesting that the normal differentiation program is disrupted in these cells. Interestingly, these lesions are only formed when both normal and mutant cells were transplanted, suggesting that FD requires the presence of normal as well as mutant osteogenic cells.
Several lines of evidence suggest that elevated cAMP may stimulate the
proliferation and inhibit the differentiation of osteogenic precursors
and therefore lead to FD. Cell culture studies demonstrate that PTH and
forskolin, which both stimulate cAMP production, inhibit osteoblast
differentiation (256, 257, 258), while treatment of cells with
a Gs antisense oligonucleotide (which
decreases Gs expression) promotes osteoblast
differentiation (259). Increased cAMP, through activation
of PKA, increases the expression of c-fos and other genes
(260). cAMP binds to the PKA regulatory subunits, which
then dissociate from the catalytic subunits. The catalytic subunits
translocate to the nucleus and phosphorylate transcription factors
[e.g., cAMP-response element binding protein (CREB)].
Phosphorylated cAMP response element binding protein (CREB) binds to
the promoter of c-fos, leading to increased expression of
the protein Fos (261). Fos overexpression in osteogenic
cells in response to increased cAMP appears to be an important common
pathway in the development of FD. Fos is one component of activator
protein-1 (AP-1) heterodimers, which themselves are
transcriptional factors. AP-1 is present at high levels in
proliferating osteoblasts, and its level declines markedly as these
cells differentiate (254). One mechanism by which high
levels of AP-1 prevent osteoblast differentiation is by inhibiting
transcription of genes that produce osteoblast-specific proteins. For
example, AP-1 directly binds to the promoter of the osteocalcin gene
and inhibits osteocalcin gene transcription (254). Fos is
overexpressed in the poorly differentiated mesenchymal cells within
human FD lesions (262), and overexpression of Fos in
transgenic mice produces bone lesions that are in many ways similar to
FD (263). In addition to their effects on osteoblast
differentiation, both cAMP and Fos stimulate the expression of IL-6.
IL-6 is overexpressed in cells within FD lesions, and increased local
levels of IL-6 stimulate the osteoclast recruitment and bone resorption
necessary for the concentric growth of these lesions (218, 264).
Other mechanisms may also play a role in the pathogenesis of FD. A recent study in three patients showed that PTH-related protein (PTHrP) is also highly overexpressed in FD lesions and suggested that increased local concentrations of PTHrP may contribute to the development of FD (265). The ß-chain of platelet-derived growth factor (PDGF-B), which stimulates fibroblast proliferation, has also been shown to be expressed at high levels in FD (266). The ability of sex steroids to increase PDGF-B expression (266, 267, 268) may account for the increased growth of FD lesions that sometimes occurs during puberty, pregnancy, or the use of oral contraceptives (269).
|
IV. Gs Loss-of-Function Mutations
|
|---|
 TopAbstractI. IntroductionII. Gs{alpha}-Signaling...III. Gs{alpha} Activating... IV. Gs{alpha} Loss-of-Function... V. Insights from Gnas...VI. GNAS1 Imprinting Mechanisms...VII. SummaryReferences |
|---|
loss-of-function
mutations lead to AHO, a syndrome characterized by skeletal and other
developmental defects. Maternal inheritance of
Gs mutations produces offspring who have both
AHO and multihormone resistance [termed pseudohypoparathyroidism type
IA (PHPIA)] while paternal inheritance of these mutations produces
offspring who have only the AHO phenotype [termed
pseudopseudohypoparathyroidism (PPHP)]. The clinical features,
diagnosis, and treatment of these disorders have been reviewed
elsewhere (270, 271), and we will briefly review the
clinical features here. We then discuss the Gs
mutations that are associated with AHO and the role of these mutations
(and the modifying effects of genomic imprinting) on the pathogenesis
of PHPIA and PPHP. Finally, we will summarize recent studies suggesting
a role for these mutations in progressive osseous heteroplasia
(POH).
A. Albright hereditary osteodystrophy (AHO)
AHO is characterized by the following signs: short stature,
brachydactyly, sc ossifications, centripetal obesity, facial features
such as depressed nasal bridge and hypertelorism, and mental deficits
or developmental delay (270, 271). The severity of the AHO
phenotype varies greatly between patients, and some patients with
inactivating Gs mutations have either few
(272) or no (273) features of the
syndrome.
Brachydactyly refers to the shortening and widening of specific long bones in the hands and feet, which in AHO most often involves the distal thumb and third, fourth, and fifth metacarpals (274, 275, 276). This pattern of involvement, which may be symmetrical or asymmetrical, is relatively specific for AHO when compared with other brachydactyly syndromes (274, 275). Brachydactyly becomes clinically obvious within the first several years of life and is characterized radiologically by premature closure and coning of the epiphyses. Ectopic intramembranous ossification can be present in any location and is generally confined to the superficial sc tissues. The skeletal manifestations of AHO are not secondary to hypocalcemia, hyperphosphatemia, or elevated circulating PTH, as they are absent in patients with primary hypoparathyroidism and present in patients with PPHP (in whom these biochemical parameters are normal).
Obesity is a common feature of AHO, being present in both PHPIA and PPHP patients. PHPIA patients have been shown to have decreased circulating FFA levels and a decreased cAMP response to ß-adrenergic stimulation in fat cell membranes (277). Another study showed that the basal and epinephrine-stimulated glycerol production rate is reduced in PHPIA patients, although another group of obese patients without a Gs defect also had decreased glycerol production rates (278). Less than 50% of AHO patients also present with mental retardation and/or delayed development (279).
B. Pseudohypoparathyroidism type IA (PHPIA)
Patients who inherit AHO from their mother present with PHPIA
(124). In addition to the AHO phenotype, the major feature
of PHPIA is the presence of multihormone resistance, which primarily
involves three hormones (PTH, TSH, and gonadotropins) that activate
Gs-coupled pathways in their target tissues. The
most clinically obvious abnormality is renal PTH resistance, which in
most cases presents as hypocalcemia, hyperphosphatemia, and elevated
circulating PTH. PTH resistance appears to develop over the first
several years of life, with hyperphosphatemia and elevated PTH
generally preceding hypocalcemia (280, 281, 282, 283). Some PHPIA
patients with PTH resistance remain eucalcemic (284, 285, 286).
In PHPIA the urinary cAMP response to administered PTH is markedly
reduced (287, 288), indicating a defect in PTH-stimulated
cAMP generation in renal proximal tubules. This leads to decreased
phosphate wasting and decreased conversion of 25-hydroxyvitamin D to
the active metabolite 1,25-dihydroxyvitamin D, two physiological
responses to PTH that are stimulated by cAMP (289, 290, 291, 292, 293, 294, 295).
Decreased circulating levels of 1,25-dihydroxyvitamin D contributes to
hypocalcemia through decreased gastrointestinal absorption of ingested
calcium and decreased mobilization of calcium from bone (284, 296, 297). Unlike patients with primary hypoparathyroidism, PHP
patients are not prone to hypercalciuria, indicating that the
anticalciuric action of PTH in the thick ascending limb is unaffected
(298).
In addition to PTH resistance, almost all PHPIA patients present with TSH resistance (299, 300). TSH levels are typically elevated at birth (283, 301, 302, 303) but may then normalize for a period of months before once again becoming elevated (283). The resistance is generally mild, with TSH levels that are only minimally elevated and thyroid hormone levels that are normal or slightly low. Consistent with a defect in TSH signaling, goiter is usually absent (lack of growth stimulation by TSH), and antithyroid antibodies are also absent (lack of autoimmune thyroid disease). Decreased stimulation of adenylyl cyclase by TSH was confirmed in thyroid membranes isolated from one PHPIA patient (304).
PHPIA patients, particularly females, present with clinical evidence of hypogonadism, which is usually manifested as delayed or incomplete sexual maturation, amenorrhea or oligomenorrhea, and/or infertility (299, 305, 306, 307, 308). As a group these women are slightly hypoestrogenic (308), but studies have not been able to consistently establish that they have increased basal or GnRH-stimulated levels of circulating gonadotropins (299, 305, 306, 307, 308). It has been proposed that PHPIA patients may have a partial resistance to gonadotropins that may allow for relatively normal follicular development, which produces adequate E production for negative feedback, but which prevents normal ovulation (308). Many, although not all, PHPIA patients also have PRL deficiency, although the underlying mechanism for this is unknown (299, 309, 310, 311, 312, 313). GH deficiency has also been reported to occur in some (306, 314), but not all (309, 315), PHPIA patients, suggesting that at least some patients may be resistant to GHRH. PHPIA patients also appear to be resistant to the lipolytic actions of epinephrine in adipocytes (278).
PHPIA patients do not show resistance to all hormones that stimulate
Gs-coupled pathways in their target tissues.
Although the plasma cAMP responses to glucagon (299, 316)
and isoproterenol (317) are reduced in these patients, the
downstream physiological response (rise in serum glucose) is normal,
indicating that the rise in cAMP is sufficient to produce a maximal
physiological response. PHPIA patients also do not appear to be
resistant to vasopressin (309, 318) or ACTH (299, 309). Several reports suggest that olfaction is impaired in
PHPIA, but not in PPHP or PHPIB, patients (319, 320, 321, 322).
Whether this reflects differences in hormonal status,
Gs levels, or other mental functions is
unknown.
C. Pseudopseudohypoparathyroidism (PPHP)
Patients who inherit AHO from their father present with PPHP
(124). PPHP patients present with the physical features of
AHO but do not have hormone resistance. In contrast to PHPIA, serum
levels of calcium, phosphate, PTH, TSH, and thyroid hormone, as well as
the urinary cAMP response to administered PTH, are normal in PPHP
patients (288). Because many of the clinical features of
AHO are nonspecific, the diagnosis of PPHP should be made only in
patients within a well documented AHO kindred or in whom a
Gs defect (decreased protein expression or
mutation, see Section IV.D) is identified. Some patients
with an AHO-like syndrome have a chromosomal deletion at 2q37
(323, 324).
D. The role of inactivating Gs mutations and
Gs imprinting in the pathogenesis of AHO, PHPIA, and
PPHP
As PHPIA patients are resistant to several hormones that activate
Gs, it seemed likely that these patients have a
defect in this signaling component. Biochemical assays that measure
hormone signaling after reconstitution of patient cell membranes with
membranes that totally lack Gs (cyc-)
demonstrated that in PHPIA functional Gs activity
is decreased by approximately 50% in membranes from various cell
types, including erythrocytes, fibroblasts, platelets, lymphocytes, and
kidney (288, 325, 326, 327, 328, 329, 330, 331, 332, 333, 334, 335). Subsequent studies showed that in
many cases this was due to decreased expression of
Gs mRNA (335, 336) and/or protein
(273, 337). Functional studies in membranes (304, 338) and whole cells (339) confirmed that these
patients have a defect in Gs signaling.
Subsequently, Gs was shown to be similarly
reduced in erythrocyte membranes isolated from PPHP patients
(288), suggesting a similar genetic defect of
Gs in these patients.
Consistent with 50% loss of Gs expression in
PHPIA and PPHP, multiple heterozygous inactivating mutations within the
Gs coding exons have been identified in these
patients (44, 57, 272, 273, 283, 340, 341, 342, 343, 344, 345, 346, 347, 348, 349, 350, 351, 352, 353, 354, 355, 356, 357, 358) (for a
compilation of mutations see Refs. 271 and
357 or the web site http://mammary.nih.gov/aho/).
These mutations are spread throughout the coding exons, with some
clustering in exons 1, 4, 5, 10, and 13 (357). One
specific 4-bp deletion has been identified in at least 13 families
(341, 344, 347, 351, 352, 353, 355, 357, 358). This probably is
not due to a founder effect, but rather to the frequent occurrence of
de novo mutation due to pausing of DNA polymerase during
replication and slipped strand mispairing (344). Two
other mutations (Met1 to Val,
Ala366 to Ser) have each been reported in two
kindreds (340, 345, 357). All other mutations have been
identified in only a single kindred. No mutations have been identified
in exon 3, possibly reflecting the fact that it is small and that it
can be spliced out and still produce a functional
Gs protein (357).
Many of the GNAS1 mutations in AHO are deletions or
insertions that produce frameshifts and premature stop codons, nonsense
mutations, or splice junction mutations (272, 283, 341, 342, 343, 344, 347, 348, 349, 351, 352, 353, 354, 355, 357, 358). Most of these mutations disrupt
expression of Gs mRNA and, consequently,
Gs protein. However, a number of missense
mutations that alter the protein sequence at one or a few amino acid
residues have also been identified (Table 2), and biochemical characterization of
some of these mutant proteins has provided important insights into the
normal function of Gs. Several of these
mutations globally disrupt protein stability due to steric effects
produced by the presence of an amino acid with a bulky hydrophobic side
chain [e.g, Ser250 to Arg
(350); Glu259 to Val, (48, 347), see Fig. 2]. Amino acid substitutions can also
destabilize the protein by leading to increased dissociation of guanine
nucleotide, as -subunits require bound guanine nucleotide for normal
thermostability (e.g., Ser250 to Arg
(350); Arg258 to Trp (44);
Ala366 to Ser (345)].
|
from receptors while a nearby mutation [deletion of
Ile382 (359)] leads to selective uncoupling from
PTH receptors and therefore selective resistance to PTH, producing a
PHPIB phenotype (Fig. 2). Substitution of the initiator methionine
produces a large, inactive form of Gs
(340). Mutation of the basic residue
Arg231 within the switch 2 region (Fig. 2) leads
to a receptor-activation defect, presumably by disrupting interactions
between this residue and acidic residues within switch 3 that stabilize
the active conformation (47, 346). Consistent with this
mechanism, mutation of the acidic residue Glu259 (Fig. 2)
in switch 3 produces a similar phenotype (48). Mutation of
the neighboring switch 3 residue Arg258 produces two major
effects. In the basal state GDP dissociates at a higher rate due to
loss of an interaction between Arg258 and the helical
domain residue Gln170, which normally forms a "lid"
over the cleft between the two domains (44, 51) (Fig. 2).
Also, mutation of this residue increases the rate of the intrinsic
GTPase, and therefore receptor activation is diminished because the
activated GTP-bound form of Gs is short lived
(51). Another mutation (Ala366 to
Ser) produces a syndrome of PHPIA plus gonadotropin-independent
precocious puberty in males (345) (Fig. 2). The mutation
leads to a subtle increase in the rate of GDP dissociation, as if the
mutant Gs was in the receptor-activated state
(360). At 37 C the mutant protein is unstable due to
increased GDP dissociation leading to loss of
Gs protein expression and the PHPIA phenotype.
However, at the slightly lower ambient temperature of the testis the
protein remains stable and is constitutively activated because GDP
dissociation is normally the rate-limiting step for G protein
activation. The resulting increased cAMP levels within the testis lead
to testotoxicosis.
Consistent with the similar Gs deficiency in
PHPIA and PPHP, identical GNAS1 mutations are present in
both PHPIA and PPHP patients within the same family (272, 283, 343, 349, 352, 353, 355), confirming that AHO is an autosomal
dominant disorder caused by GNAS1 mutations. For many years
it was unclear why some patients develop hormone resistance (PHPIA)
while other patients do not (PPHP). The first clue in explaining this
discrepancy was provided by the observation that maternal inheritance
of AHO produces offspring with PHPIA, while paternal inheritance of AHO
produces offspring with PPHP (124). Therefore, whether or
not an individual develops PHPIA or PPHP is dependent on the sex of the
affected parent and not on whether the affected parent has PHPIA or
PPHP. This parent-of-origin-specific inheritance pattern, which has
been confirmed in other studies (272, 283, 343, 349, 352, 353, 355), strongly suggests that GNAS1 is imprinted.
An imprinting model predicts that in specific hormone target tissues
(e.g., renal proximal tubules, the major site of PTH action
in the kidney), Gs is primarily expressed from
the maternal allele due to suppression (imprinting) of the paternal
allele (Fig. 4). If an inactivating
mutation is inherited from the mother, then Gs
expression is lost due to mutation of the active maternal allele,
leading to loss of hormone signaling (PHPIA). In contrast, a mutation
on the inactive paternal allele would not affect
Gs expression or hormone signaling (PPHP).
Consistent with this model, the urinary cAMP response to administered
PTH is markedly reduced in PHPIA patients but is normal in PPHP
patients (287, 288). Assuming this model is correct, the
imprinting of Gs would have to be limited to
specific tissues, as Gs is biallelically
expressed in several human tissues (77, 79, 126). Other
genes are also imprinted in a tissue-specific manner
(361, 362, 363, 364). Biallelic expression of
Gs would explain why in several tissues
Gs expression is similarly reduced by
approximately 50% in both PHPIA and PPHP patients (288)
(Fig. 4). As in humans, a maternal Gs mutation
in mice also produces renal PTH resistance while the same mutation in
the paternal allele does not, and molecular studies confirm that
Gs is paternally imprinted (suppressed) in a
tissue-specific manner (125) (see Section V).
Gs has also been shown to be paternally
imprinted in human pituitary glands (127), although the
imprinting status of Gs in human renal
proximal tubules has not as yet been assessed.
|
imprinting may explain why PHPIA patients fail to demonstrate
resistance to other hormones that also activate
Gs in their target tissues (e.g.,
ACTH, vasopressin). If Gs is not imprinted in
their respective target tissues (e.g., adrenal cortex, renal
collecting ducts), mutations should reduce Gs
expression by only 50%. Partial loss of Gs
may fail to produce hormone resistance, either because
Gs is not rate limiting for hormone-stimulated
cAMP, or because the reduced cAMP levels are still above the threshold
required to elicit the downstream physiological responses (316, 365, 366). Lack of Gs imprinting in the
distal nephron (125, 367) may explain why the
anticalciuric effect of PTH in the thick ascending limb is maintained
in PHPIA patients (298).
Although the multihormone resistance associated with AHO is very likely
to be the direct consequence of Gs deficiency,
the mechanisms that lead to the skeletal, developmental, and
neurological manifestations of AHO are less well defined. Due to their
opposite imprinting (Fig. 3), NESP55 expression will be disrupted only
by mutations within exons 2–13 on the maternal allele that affect mRNA
expression, while XLs expression will be disrupted only by mutations
on the paternal allele that affect mRNA expression or protein function.
Because the AHO phenotype is similar in PHPIA patients who inherit the
disease maternally and PPHP patients who inherit the disease
paternally, it appears unlikely that defects in either NESP55 or XLs
are involved in the development of AHO. Moreover, loss of NESP55
expression in PHPIB patients is not associated with any of the
manifestations of AHO (89). The AHO phenotype probably
results from haploinsufficiency of Gs in many
tissues in both PHPIA and PPHP patients (Fig. 4), although the exact
mechanisms involved remain to be defined.
Studies in patients with FD suggest that activating
Gs mutations inhibit osteoblast
differentiation (see Section III.C). Conversely, decreased
Gs signaling may promote osteoblast
differentiation, resulting in ectopic ossification in AHO patients (as
well as in POH patients, see below). Bones affected with brachydactyly
have premature closure of growth plates. Bone lengthening occurs at the
growth plates through endochondral ossification, a process in which
proliferating chondrocytes differentiate into hypertrophic
chondrocytes, which then become ossified. PTHrP, which activates
Gs by binding to the PTH/PTHrP (PTH1) receptor, acts in a
paracrine manner to inhibit the transition to hypertrophic chondrocytes
(368), and inactivating mutations in the PTHrP
(369) or PTH1 receptor (370, 371) genes
produce skeletal dysplasias caused by accelerated chondrocyte
differentiation within the growth plate. A small number of
Gnas knockout mice also present with growth plate
abnormalities (Ref. 125 and S. Yu and L. S.
Weinstein, unpublished data) and in one recent study growth plate
chondrocytes with the same Gnas mutation differentiated more
rapidly than wild-type chondrocytes (372).
Agents that stimulate lipolysis in adipocytes, such as norepinephrine
released by sympathetic nerves or epinephrine secreted by the adrenal
medulla, activate Gs and raise intracellular cAMP. One
mechanism by which increased cAMP stimulates lipolysis is through
phosphorylation of perilipin by PKA (373). In the
nonphosphorylated state, perilipin is localized to the surface of
intracellular lipid droplets, and upon phosphorylation perilipin moves
away from the lipid droplet, allowing hormone-sensitive lipase to have
access to the lipid substrate. Mice lacking perilipin
(374) or a PKA-regulatory subunit (resulting in increased
PKA activity, Ref. 375) are lean. In contrast, mice with
an increased ratio of 2 to ß-adrenergic receptors in adipocytes
(which lowers the ability of catecholamines to raise intracellular
cAMP) are prone to obesity (376). In PHPIA patients both
the cAMP (277) and lipolytic (277, 278)
responses to catecholamines are decreased, presumably due to low levels
of Gs in adipocytes. Results in
Gnas knockout mice suggest that obesity may result from
decreased metabolic activation of adipose tissue caused by decreased
activity of sympathetic nerves (128). In one study, PHPIA
patients had low plasma levels of norepinephrine and normal epinephrine
levels, consistent with decreased sympathetic nerve activity
(278). Further studies will be required to determine
whether the decreased lipolysis rate in PHPIA is due to decreased white
adipose tissue responsiveness, decreased sympathetic stimulation, or
both. ß-Adrenergic-Gs pathways also stimulate the
expression of uncoupling protein 1 and thermogenesis in brown adipose
tissue (377). Although Gs
deficiency in brown adipose tissue would be predicted to promote the
development of obesity by decreasing energy expenditure, there is
little evidence that brown adipose tissue has a major role in the
maintenance of energy balance in humans. In addition to its effects on
metabolism, Gs deficiency may also promote
adipocyte differentiation. Treatment of 3T3-L1 preadipocytes with
antisense Gs oligonucleotides promoted
adipocyte differentiation in a cAMP-independent manner, while treatment
with cholera toxin prevented their differentiation (378).
In another study, activating Gs mutations
prevented the differentiation of transplanted marrow stromal cells to
adipocytes (255).
cAMP is known to be important for learning and memory, and mutation of
adenylyl cyclase or PKA produces learning and memory defects in
Drosophila (379) and mice
(380, 381, 382). Exactly how Gs
deficiency might lead to the mental deficits in AHO and why these
deficits are present in only a subset of AHO patients remain to be
determined.
E. Progressive osseous heteroplasia (POH)
POH is a congenital disorder of ectopic bone formation that was
first described in 1994 (383). POH is characterized by the
development of ossifications within the dermis and sc fat that grow
laterally and coalesce to form plaques and then subsequently grow
inwardly to involve deep connective tissues and skeletal muscle
(384). Ectopic bone formation in POH is primarily by
intramembranous, rather than endochondral, ossification. This disorder
affects both males and females and in some cases is inherited in an
autosomal dominant pattern. POH is easily distinguishable from
fibrodysplasia ossificans progressiva, a disorder associated with
heterotopic endochondral ossification and overexpression of bone
morphogenetic protein 4, and is generally not associated with the other
characteristic features of AHO. Unlike the ossification in POH, the
ossification in AHO is generally limited to the superficial sc
tissues.
Recently, heterozygous inactivating GNAS1 mutations or
reduced Gs expression have been shown to be
associated with POH in three patients, two of whom show other signs of
AHO. In the first case, a 4-bp deletion in exon 7 which is found in
several AHO patients was present in a girl with plate-like osteoma
cutis involving the face, scalp, and extraocular muscle (a variant of
POH) and other areas of heterotopic ossification in the absence of
other features of AHO or hormone resistance (385). In the
second case, a nonsense mutation within exon 1 at the codon encoding
Gln12 was present in a girl with POH and mild
brachydactyly, but without other signs of AHO or hormone resistance
(386). In the third case with POH and PHPIA,
Gs expression was decreased in erythrocyte
membranes, although a specific GNAS1 mutation was not
identified (386). It remains to be determined whether more
typical cases of POH without AHO are also associated with
GNAS1 mutations. The fact that the 4-bp deletion in exon 7
has been identified in both AHO and POH patients makes it unlikely that
POH is due to a specific subset of mutations that more severely affect
Gs function. It is likely that the severity
and extent of ectopic calcifications in patients with
Gs mutations are determined by other unrelated
"modifier" genes.
The presence of activating Gs mutations in FD,
which is characterized by a block in osteoblast differentiation, and
inactivating Gs mutations in AHO and POH,
which are associated with intramembranous ossifications in ectopic
locations, provides in vivo evidence that
Gs pathways negatively regulate osteoblast
differentiation. Cell culture studies provide evidence that increased
cAMP inhibits osteoblast differentiation (256, 257, 258) and
decreased Gs expression promotes osteoblast
differentiation (259). In the patient with plate-like
osteoma cutis and a Gs exon 7 mutation, an
osteoblast-specific isoform of the osteogenic transcription factor
Cbfa1/RUNX2 was expressed in dermal fibroblasts, suggesting that they
may be more likely to differentiate into an osteoblastic phenotype
(385).
|
V. Insights from Gnas Knockout Mice |
|---|
Gnas is located within the distal chromosome 2 imprinted
region (69, 70). To test whether Gnas is
involved in the UPD phenotypes, we created a Gnas knockout
mouse by targeted mutagenesis. The resultant mutant Gnas
allele had an insertion in Gnas exon 2, an exon that is
included in all Gnas transcripts (Ref. 125 , see Fig. 3).
Double knockouts (-/-) died in early gestation, indicating that the
gene is required for normal development. Heterozygotes with disruption
of the maternal (m-/+) or paternal (+/p-) allele had distinct
phenotypes, providing strong evidence that Gnas is
imprinted. M-/+ mice are larger than normal at birth with flat, broad
bodies and sc edema that resolves soon after birth. Most m-/+ mice
develop neurological abnormalities and die anywhere from 1–3 wk after
birth. In contrast, +/p- mice are born small with narrow bodies; they
fail to suckle, and most die several hours after birth, most likely
from hypoglycemia. Interestingly, the abnormal phenotypes of m-/+ and
+/p- mice are almost identical to those described for paternal and
maternal UPD mice, respectively (104, 125) (Fig. 5A). A mutation produced by radiation
mutagenesis, which maps close to Gnas, also produces similar
phenotypes upon maternal or paternal transmission
(388).
|
The similar phenotypes of Gnas knockout and distal
chromosome 2 UPD mice suggest that m-/+ and +/p- phenotypes are due
to loss of maternal and paternal-specific Gnas gene
products, respectively. While some aspects of the m-/+ phenotype could
theoretically result from loss of NESP55 expression, we speculate that
NESP55 does not play a significant role in the development of the m-/+
phenotype. The exon 2 disruption in Gnas knockout mice does
not disrupt the NESP55 coding region (Fig. 3), and RT-PCR experiments
indicate that NESP55 mRNA expression is not lost in m-/+ mice (S. Yu
and L. S. Weinstein, unpublished results). Moreover, loss of
NESP55 expression in humans does not produce an obvious phenotype
(89). Loss of XLs expression could contribute to the
+/p- phenotype. However, mutation of the paternal allele in PPHP
patients, which should disrupt XLs expression, does not seem to
produce an obvious phenotype that is unique to PPHP. Differences in
Gs expression between m-/+ and +/p- mice due
to the tissue-specific imprinting of Gs are
likely to contribute to the phenotypic differences. It is also possible
that there are additional Gnas gene products that are at
present unidentified. Specific knockouts of individual Gnas
gene products will be useful in determining to what extent each
contributes to the Gnas knockout phenotypes.
In many respects the m-/+ and +/p- mouse phenotypes do not correlate to the respective human PHPIA and PPHP phenotypes (125, 128). These differences might reflect species-specific differences in the importance of Gnas gene products or in other physiological or environmental factors.
A. Tissue-specific imprinting of Gs
M-/+ mice have decreased circulating levels of ionized calcium
and increased levels of phosphate and PTH, consistent with the presence
of PTH resistance, while calcium, phosphate, and PTH levels are normal
in +/p- mice (125). Moreover, PTH-stimulated cAMP
generation in renal proximal tubules is reduced by approximately
70% in m-/+ mice but is unaffected in +/p- mice. Therefore, in both
human and mouse, mutation of the maternal allele (PHPIA patients, m-/+
mice) is associated with PTH resistance while mutation of the paternal
allele (PPHP patients, +/p- mice) is not associated with PTH
resistance. Consistent with these biochemical observations,
Gs mRNA and protein expression in renal
proximal tubules is significantly reduced in m-/+ mice but is
unaffected in +/p- mice (125) (Fig. 4). These results
confirm that Gs is imprinted in mouse proximal
tubules, with expression mostly derived from the maternal allele. PTH
resistance in m-/+ mice (and presumably in PHPIA patients) results
from markedly reduced Gs expression due to
mutation of the active maternal allele. In contrast, mutation of the
inactive paternal allele in +/p- mice (and presumably PPHP patients)
has little effect on Gs expression or PTH
signaling. The presence of some residual Gs
expression and PTH-stimulated cAMP production in m-/+ mice suggests
that Gs expression from the paternal allele is
not fully suppressed.
In one study Gs mRNA was shown to be more
abundant in the glomeruli of mice with paternal UPD of the distal
chromosome 2 imprinted region and less abundant in mice with maternal
UPD of the same region, suggesting that the paternal, rather than the
maternal, allele is preferentially expressed in glomeruli
(389). No similar differences in glomerular
Gs expression were observed in Gnas
knockout mice (125). The apparent preponderance of
glomerular Gs expression from the paternal
allele in UPD mice might reflect differences in kidney maturation, as
the UPD mice were studied during late gestation, and the rate of kidney
maturation is affected by mutation of Gnas
(125). It is also possible that the increased signal in
paternal UPD mice reflects increased expression of an alternative
Gnas gene product (e.g., XLs), as a probe
common to all Gnas gene products was used in this study.
Interestingly, the imprinting of Gs in the
kidney appears limited to the renal proximal tubules, as there is no
evidence of imprinting in the inner medulla (composed primarily of
collecting ducts) and outer medulla (consisting primarily of thick
ascending limbs). In both of these regions Gs
expression is similarly reduced by about 50% in m-/+ and +/p- mice,
respectively (125, 367) (Fig. 4).
Gs is also not imprinted in other tissues
(e.g., lung, skeletal muscle). Gs
imprinting in humans is also tissue specific, as
Gs is biallelically expressed in several human
tissues (77, 79, 126) and Gs
expression in various tissues is similarly reduced in both PHPIA and
PPHP patients (288).
Gs imprinting is probably not limited to
the renal proximal tubules. For example, Gs
expression is markedly reduced in brown and white adipose tissue of
m-/+, but not +/p-, mice (125). Although this may be due
to maternal-specific expression of Gs in
adipose tissue, it is also possible that these differences in
Gs expression are secondary to differences in
metabolic activation of adipose tissue in m-/+ and +/p- mice,
respectively (128) (see Section V.C).
B. Role of Gs in renal function
Several hormones, including PTH, vasopressin, calcitonin, and
glucagon, have receptors within specific segments of the nephron that
couple to Gs to produce their physiological
effects in the kidney. The role of Gs defects
in renal function has been recently reviewed (365).
Because imprinting of Gs in the nephron
appears to be confined to the proximal tubules, the major renal
abnormality in m-/+ mice and in PHPIA patients is renal PTH resistance
in the proximal tubule, leading to decreased phosphate reabsorption and
synthesis of 1,25-dihydroxyvitamin D, as summarized in Section
III. PTH also acts on the thick ascending limb where it stimulates
calcium reabsorption, and for this reason patients with primary
hypoparathyroidism who lack PTH are prone to hypercalciuria after
treatment with calcium and vitamin D. In contrast, PHPIA patients
generally do not develop hypercalciuria (298).
Gs is not imprinted in the thick ascending
limb, and therefore Gs levels in the thick
ascending limb might suffice to maintain adequate calcium reabsorption.
Alternatively, the calcium reabsorption response may not be dependent
on cAMP, but rather on other PTH-stimulated effectors (e.g.,
Gq-stimulated intracellular calcium).
Consistent with approximately 50% loss of Gs
expression in the thick ascending limb of m-/+ and +/p- mice, both
groups of mice have an approximately 50% loss of expression of the
cAMP-regulated Na-K-2Cl cotransporter (BSC1) in the thick ascending
limb (367). As this transporter is necessary for the
generation of high solute concentrations in the renal medulla,
decreased expression of this protein would be expected to result in
decreased urine concentration in response to acute vasopressin
administration. Urine concentration 1 h after an ip bolus of
vasopressin (which produces maximal water permeability in the
collecting ducts) was decreased by 28% in m-/+ mice
(367). Levels of adenylyl cyclase 6 are also decreased in
the thick ascending limb of mutant mice, and this may also lead to
lower cAMP levels (367). In contrast, the acute urinary
concentrating response to vasopressin was normal in three PHPIA
patients (318). After chronic water deprivation (which
leads to maximal circulating vasopressin), mutant and wild-type mice
increase BSC1 expression (367) and urine concentration
(125) to similar levels, suggesting that in both groups of
animals vasopressin can produce maximal cAMP levels in the thick
ascending limb that exceed those required for maximal BSC1
expression.
Vasopressin stimulates water reabsorption in the collecting ducts by
two mechanisms: chronically by increasing expression of the water
channels aquaporin 2 and 3 (AQP2, AQP3) and acutely by stimulating the
translocation of AQP2 to the apical membrane of tubule cells in the
collecting ducts (390, 391). Although AQP2 expression is
decreased by approximately 50% in the outer medulla (cortical
collecting ducts) of both m-/+ and +/p- mice, AQP2 is expressed at
normal levels in the inner medulla (inner medullary collecting ducts)
in mutant animals (365, 367). Therefore, cAMP levels
appear to be limiting for AQP2 expression in the collecting but not the
inner medullary collecting ducts. Expression of the
vasopressin-regulated urea transporter and adenylyl cyclase 6 are also
normal in the inner medullary collecting ducts of mutant mice
(367). Although AQP2 translocation has not been directly
examined in Gnas knockout mice, vasopressin-stimulated cAMP
levels greatly exceed those normally required for maximal water
permeability (392), and therefore it is likely that these
responses are well maintained in the collecting ducts in
Gnas knockout mice. Based on the lack of
Gs imprinting in collecting ducts and the
excess capacity built into the water reabsorption machinery in
collecting ducts, it is not surprising that neither Gnas
knockout mice (125, 367) nor PHPIA patients (309, 318) have any of the clinical features of vasopressin
resistance.
C. Role of Gs in energy and glucose metabolism
One surprising phenotypic difference between m-/+ and +/p- mice
is the opposite effects on energy metabolism (128). M-/+
mice become obese and are hypometabolic and hypoactive while +/p- mice
have markedly reduced lipid accumulation in white and brown adipose
tissue and are hypermetabolic and hyperactive. The opposite effects on
white and brown adipose tissue are obvious by 2 d after birth
(Fig. 5B). These effects are not due to altered thyroid hormone status,
food intake, or leptin regulation, but rather appear to result from
differences in metabolic activity. Similar opposite effects on weight
accumulation are also present in mice with maternal and paternal
inheritance of a radiation-induced mutation that maps near
Gnas (388).
In +/p- mice the sensitivity of adipose tissue to sympathetic nervous
system stimulation should be normal, as Gs
expression in this tissue is unaffected (125). In m-/+
mice Gs expression in adipose tissue is
significantly reduced presumably due to imprinting (125),
and one might expect adipose tissue in these mice to be resistant to
sympathetic nerve stimulation. However, m-/+ mice have normal
metabolic responsiveness to a ß3-adrenergic
agonist, an agent that specifically stimulates adipose tissue in
vivo (128). Urinary excretion of norepinephrine is
decreased in m-/+ mice and increased in +/p- mice, suggesting that
sympathetic activity is decreased and increased in m-/+ and +/p-
mice, respectively (128). These changes could produce the
opposite effects on adipose tissue observed in these two groups of
mice.
The obesity in m-/+ mice mimics the obesity typically present in PHPIA patients. PHPIA patients have decreased circulating FFA levels (277) and decreased rates of basal and epinephrine-stimulated glycerol production (278). Interestingly, three PHPIA patients were reported to have low circulating levels of norepinephrine (278), which raises the possibility that sympathetic activity may also be reduced in PHPIA patients. The decreased fat mass observed in +/p- mice does not mimic the comparable human model, as PPHP patients tend to be obese rather than lean. The disparate findings between +/p- mice and PPHP patients might reflect species-specific differences in the role of brown adipose tissue in energy balance, the importance of specific Gnas gene products, or social and environmental factors.
Both obese m-/+ and lean +/p- mice have increased sensitivity to
insulin in vivo and increased insulin-stimulated glucose
uptake in skeletal muscle (393). Obesity is generally
associated with insulin resistance, so the presence of increased
insulin sensitivity in obese m-/+ mice is noteworthy.
Gs expression in skeletal muscle is decreased
to a similar extent in m-/+ and +/p- mice [presumably
Gs is not imprinted in this tissue
(128)], and this may explain the similar changes in
insulin sensitivity in muscle observed in these two groups of mice.
These findings suggest that Gs is a negative
regulator of insulin action. Several agents that counterregulate
insulin action (e.g., glucagon, catecholamines) activate
Gs pathways, and many in vivo and
in vitro studies indicate that Gs and
cAMP negatively regulate insulin-stimulated glucose uptake
(394, 395, 396, 397, 398, 399, 400). Gs may also directly
inhibit the rate of incorporation of the insulinsensitive glucose
transporter GLUT4 into plasma membranes in response to insulin
(401).
|
VI. GNAS1 Imprinting Mechanisms and Defects |
|---|
|
TopAbstractI. IntroductionII. Gs{alpha}-Signaling...III. Gs{alpha} Activating...IV. Gs{alpha} Loss-of-Function...V. Insights from Gnas...VI. GNAS1 Imprinting Mechanisms...VII. SummaryReferences |
|---|
, or adenylyl cyclase.
Although the PTH1 receptor gene appeared to be a good candidate gene
for PHPIB (403), several lines of evidence, including
mutation screening (404, 405), mouse knockout
(370) and human disease (371) models, and
mapping studies (402, 406), have ruled out the receptor as
the site of the defect in PHPIB. In four PHPIB kindreds, the disease
was mapped to 20q13, in the vicinity of GNAS1, suggesting
that PHPIB, like PHPIA, is due to a genetic defect involving
GNAS1 (402). However, Gs
function is unaffected in erythrocyte membranes from PHPIB patients
(299, 403), ruling out mutations within exons 1–13 that
disrupt Gs.
UPD or so-called "imprinting" mutations, in which both alleles of
an imprinted gene have the same parental imprinting pattern even in the
absence of UPD, can both produce abnormal phenotypes through the over-
or underexpression of imprinted gene products (101, 109, 139, 407, 408, 409). If both GNAS1 alleles have a
paternal-specific imprinting pattern, then Gs
expression in renal proximal tubules will be markedly reduced due to
lack of an "active maternal" allele, resulting in renal PTH
resistance. In contrast, abnormal GNAS1 imprinting will not
affect Gs expression in most other tissues in
which Gs is normally equally expressed from
the maternal and paternal allele. This model would account for the
combination of renal PTH resistance and normal erythrocyte
Gs expression that is present in PHPIB
patients. Normal Gs expression in the vast
majority of tissues in which Gs is
biallelically expressed might also explain why these patients have no
features of the AHO phenotype (Fig. 6).
|
. In 18 of 18 PHPIB patient
blood samples studied to date (Ref. 89 and J. Liu and
L. S. Weinstein, unpublished data) the exon 1A DMR was
unmethylated on both alleles, and in several informative patients, exon
1A mRNA transcripts were biallelically expressed (Fig. 6). These
results are consistent with both alleles having a paternal-specific
imprinting pattern at the exon 1A DMR (unmethylated, transcriptionally
active). This defect was present in both sporadic and familial cases
and also in patients who initially presented with severe skeletal
changes resulting from excess circulating PTH (so-called
pseudohypohyperparathyroidism). In contrast, the imprinted
NESP55 and XLs promoter regions were abnormally imprinted (with a
paternal-specific imprinting pattern in both alleles) in only 5 of 13
and 2 of 13 PHPIB patients, respectively (89). This makes
it unlikely that either the NESP55 or XLs promoters or their gene
products are involved in the pathogenesis of PHPIB or in the
tissue-specific imprinting of Gs.
Interestingly, loss of NESP55 expression due to methylation of both
alleles in a subset of PHPIB patients is not associated with an obvious
phenotype (89). In summary, these results show that PHPIB
results from abnormal imprinting of the exon 1A DMR. Methylation
analysis of this region is a useful diagnostic tool in the evaluation
of patients with PTH resistance.
Paternal UPD was ruled out in 12 of 13 patients (89).
However, in one patient paternal UPD of chromosome 20 was reported to
be associated with PTH resistance in addition to other abnormalities
that are possibly due to over- or underexpression of other imprinted
genes on chromosome 20 (410). In the majority of PHPIB
patients there appears to be a failure to switch from the paternal to
the maternal imprinting pattern in the oocyte, which would explain why
PTH resistance in familial PHPIB is only observed after maternal
transmission (402). Mapping of the disease to 20q13
(402) suggests that, at least in familial cases, there is
an underlying genetic defect within GNAS1 that leads to
abnormal imprinting, although such a defect remains to be identified.
Recently, three siblings with PHPIB were shown to have a
Gs mutation that deletes an amino acid
(Ile382) in the carboxyl-terminal tail (Fig. 2). This
mutant Gs protein is selectively uncoupled
from the PTH1 receptor (359). Consistent with the
imprinting model, transmission of the mutation from the maternal
grandfather to the proband’s mother did not lead to PTH resistance,
while maternal transmission of the mutation led to PTH resistance in
all three siblings with the mutation.
As discussed in Section V, maternal, but not paternal,
transmission of inactivating Gs mutations also
leads to partial TSH resistance, suggesting that
Gs is also imprinted in thyroid. Based on the
relatively small increases in circulating TSH and mild hypothyroidism
in PHPIA patients, the imprinting in the thyroid is probably only
partial, and therefore some paternal-specific expression of
Gs is maintained. In PHPIB patients two
alleles with a paternal-specific imprint might also reduce
Gs expression in the thyroid, although to a
lesser extent than in PHPIA patients, in whom the more active maternal
allele is mutated, leaving a single paternal allele. Although the
initial paper examining thyroid function in a small number of PHPIB
patients showed no evidence for TSH resistance (299), we
observe that some PHPIB patients with a GNAS1 imprinting
defect have borderline or mildly elevated levels of circulating TSH,
suggesting that at least some PHPIB patients have slight resistance to
TSH (L. S. Weinstein and J. Liu, unpublished data). TSH resistance
has also been reported in one patient with paternal UPD of 20q
(410).
B. Potential GNAS1 imprinting mechanisms
Several important questions need to be answered to gain a better
understanding of how imprinting of GNAS1 is maintained and
established, and these answers should provide more general insights
into how genes become imprinted. First, what are the
cis-acting DNA elements that target the exon 1A DMR for
methylation in the oocyte, and why does this region not get methylated
in PHPIB patients? Two DNA sequence elements have been identified that
are required for de novo methylation of the Igf2r
gene methylation imprint mark in female germ cells (411).
Other than this study, little is known about what signals are required
to establish the differential methylation of imprinted genes in male
and female gametes. The mapping of familial PHPIB to the vicinity of
GNAS1 suggests that these patients harbor mutations within
the GNAS1 locus that prevent the methylation of the exon 1A
DMR in oocytes. Identification of such mutations may allow us to define
regions of the gene that are critical for the establishment of
imprinting.
The second major question is how do the various imprinted regions of
GNAS1 regulate each other? Is there one "imprinting
center" that is required to establish imprinting throughout the
GNAS1 locus (as is the case for other imprinted gene
regions) or is imprinting at the NESP55, XLs, and exon 1A promoter
regions established independently of one another? While we do not have
the answers to these questions, the imprinting patterns in PHPIB
patients suggest that imprinting of these regions may be coregulated in
some, but not all, cases (89). The exon 1A DMR is a
methylation imprint mark and therefore is a candidate for being a
GNAS1 imprinting center (74). However, in most
PHPIB patients the NESP55 and XLs promoters imprint normally even
when imprinting at the exon 1A DMR has not been established
(89). Imprinting of the NESP55 promoter in mouse is
established later during postimplantation development, and therefore
this region is not likely to be an imprinting center (74).
Further studies are necessary to determine the role of the NESP
antisense transcript in the imprinting mechanism and the stage in
development in which imprinting of XLs is established.
Finally, what are the mechanisms by which Gs
is imprinted in a tissue-specific manner? Although the mechanisms of
Gs imprinting have yet to be defined, recent
studies provide some testable models. Gs
imprinting does not involve methylation of its promoter. Rather, the
close proximity of the exon 1A DMR to the Gs
promoter and its abnormal imprinting in virtually all PHPIB patients
strongly suggest that this region is important for the allele-specific
expression of Gs. Because
Gs imprinting is limited to specific tissues
while the exon 1A DMR is differentially methylated in all tissues, the
imprinting mechanism must also involve another tissue-specific factor.
Reciprocal regulation of the exon 1A and Gs
promoters on the paternal allele (promoter competition model, Fig. 7A) is unlikely, because in mouse exon 1A
mRNA expression is not limited to the tissues where
Gs is imprinted, as this model would predict
(74). Alternatively, this region may contain a boundary
element that insulates the Gs promoter from an
upstream enhancer on the paternal allele (insulator model, Fig. 7, B
and C). Methylation of the boundary element on the maternal allele
would prevent binding of an insulator protein (e.g., CTCF)
and would therefore allow transcription from the maternal
Gs promoter to be stimulated by the upstream
enhancer. This is precisely the mechanism by which the H19
DMR controls the imprinting of Igf2, except that the DMR and
enhancers are downstream rather than upstream of the Igf2
promoter (110, 117, 118). In this model, either the
upstream enhancers are tissue specific or the boundary elements
are tissue specific (perhaps by utilizing a tissuespecific, rather
than a ubiquitously expressed, insulator protein). Finally, the
exon 1A DMR may contain a silencer element that can bind a
tissue-specific repressor on the paternal allele, but which can not
bind the repressor on the maternal allele because the binding site is
methylated (silencer model, Fig. 7D). Recent studies suggest that
tissue-specific silencers may be important in the imprinting of both
Igf2 and H19 (412, 413, 414, 415).
|
|
VII. Summary |
|---|
mutations
lead to opposite effects on cAMP generation, which most likely explain
the opposite effects on endocrine function and osteoblast development.
For example, activating mutations lead to the apparent stimulation of
peripheral endocrine glands even in the absence of trophic hormone
(e.g., pituitary tumors, MAS) while inactivating mutations
lead to hormone resistance (PHPIA). Also, activating
Gs mutations inhibit osteoblast
differentiation, leading to FD, while inactivating mutations appear to
promote osteoblast differentiation, leading to ectopic ossifications in
AHO and POH. It remains to be determined whether the other nonendocrine
manifestations of MAS and AHO result from altered cAMP generation or
are due to abnormal regulation of other Gs
effectors or perhaps other GNAS1 gene products.
Activating Gs mutations are somatic, and the
specific manifestations in a given individual are primarily determined
by the distribution of mutant-bearing cells. Patients with a widespread
distribution of such cells will have multiple manifestations (MAS),
while those with a more limited distribution will have only one or two
manifestations (endocrine tumor, FD). As inactivating
Gs mutations are generally in the germline,
other factors must determine the specific manifestations in a given
patient. Presumably, differences in other nonlinked genes or in
environmental factors contribute to the variable severity of the AHO
phenotype between individuals and the development of a more aggressive
form of ectopic ossification (POH) in a few patients. Another source of
variability is the fact that Gs is imprinted,
which will lead to different phenotypes depending on parental
inheritance.
Gs is biallelically expressed (not imprinted)
in most tissues, but in a small number of tissues, such as the renal
proximal tubules, is presumed to be expressed primarily from the
maternal allele. A region upstream of the Gs
promoter (the exon 1A DMR) is methylated only on the maternal allele in
all somatic tissues. We propose that in some tissues, such as the renal
proximal tubules, the unmethylated exon 1A DMR inhibits the expression
of Gs from the paternal allele, while in most
other tissues, Gs expression is not influenced
by the exon 1A DMR (Fig. 6, Normal). Based on this model, in renal
proximal tubules, inactivating mutations on the maternal allele should
markedly reduce Gs expression and lead to PTH
resistance (PHPIA) while mutation of the paternal allele should have
little effect on Gs expression or PTH
signaling (PPHP). In most other tissues in which
Gs is biallelically expressed, both maternal
and paternal mutations should produce a similar approximately 50% loss
of Gs expression, as is observed in both PHPIA
and PPHP. Partial deficiency of Gs in these
tissues may underlie the AHO phenotype that is present in both PHPIA
and PPHP patients. In PHPIB patients there is no inactivating mutation
of the Gs coding region, but rather an
imprinting defect producing a paternal-specific imprinting pattern
within the exon 1A DMR on both alleles. This should also result in
reduced Gs expression in renal proximal
tubules (and thus PTH resistance) but does not affect
Gs expression in most other tissues, which may
explain the lack of an AHO phenotype in these patients. Conceptually,
AHO appears to be one phenotype that is due to haploinsufficiency of
Gs and is not affected by imprinting, while
renal PTH resistance is an independent phenotype that is subject to
imprinting effects (maternal mutation in PHPIA or abnormal imprinting
in PHPIB) (Fig. 6).
|
Acknowledgments |
|---|
model in Fig. 2. This work was supported by NIDDK Division of Intramural Research.
|
Footnotes |
|---|
, Gs
-subunit; MAS, McCune-Albright syndrome; MOFD, monoostotic fibrous
dysplasia; PDGF-B, ß-chain of platelet-derived growth factor; PHPIA,
pseudohypoparathyroidism type IA; PHPIB, pseudohypoparathyroidism type
IB; POFD, polyostotic fibrous dysplasia; PPHP,
pseudopseudohypoparathyroidism; POH, progressive osseous heteroplasia;
PTHrP, PTH-related protein; RGS, regulator of G protein signaling; UPD,
uniparental disomy. |
References |
|---|
in the pathogenesis of
Albright hereditary osteodystrophy. Trends Endocrinol Metab 10:81–85[CrossRef][Medline]
mediates epidermal growth factor-elicited
stimulation of rat cardiac adenylate cyclase. J Biol Chem 265:21317–21322-subunit of Gs. J Biol
Chem 272:5413–5420s and
inhibition of NADPH oxidase by Gßs. J
Biol Chem 275:35920–35925 by the epidermal
growth factor receptor involves phosphorylation. J Biol Chem 271:6947–6951 subunit of the G protein Gs activate
both adenylyl cyclase and calcium channels. Science 243:804–807s subunit incorporates
[3H]palmitic acid and mutation of cysteine-3
prevents this modification. Biochemistry 32:8057–8061[CrossRef][Medline]
subunits
are palmitoylated. Proc Natl Acad Sci USA 90:3675–3679-subunits in rat
pancreatic endocrine cells. J Histochem Cytochem 47:289–302
subunit of Gs in intact cells alters its
abundance, rate of degradation, and membrane avidity. J Cell Biol 119:1297–1307. Cell 77:1063–1070[CrossRef][Medline]
. Mol Biol Cell 7:1225–1233[Abstract]
subunits. Proc Natl Acad Sci USA 96:412–417 the ability to interact with regulators of
G protein signaling. Biochemistry 37:13776–13780[CrossRef][Medline]
. Science 278:1943–1947·GTPS. Science 278:1907–1916 complexed with GTPS. Nature 366:654–663[CrossRef][Medline]
-subunit of a
heterotrimeric G protein. Nature 369:621–628[CrossRef][Medline]
-GDP-AlF4-. Nature 372:276–279[CrossRef][Medline]
1 and the mechanism of GTP hydrolysis.
Science 265:1405–14121ß12.
Cell 83:1047–1058[CrossRef][Medline]
1. Science 270:954–960s signal transduction protein. Proc Natl
Acad Sci USA 83:8893–8897 gene. Proc Natl Acad
Sci USA 85:2081–2085 in a patient with Albright hereditary
osteodystrophy impairs GDP binding and receptor activation. J Biol
Chem 273:23976–23983 subunit is essential for
target activation. J Biol Chem 272:21673–21676 mutant. Proc
Natl Acad Sci USA 94:5656–5661 demonstrates the importance of interactions
between switches 2 and 3 for activation. J Biol Chem 274:4977–4984 impair receptor-mediated
activation by altering receptor and guanine nucleotide binding. J
Biol Chem 273:15053–15060: evidence
for intramolecular signal transduction. Mol Pharmacol 53:981–990-subunit with impaired receptor-mediated activation because of
elevated GTPase activity. Proc Natl Acad Sci USA 96:4268–4272 chain
of Gs and stimulate adenylyl cyclase in human
pituitary tumours. Nature 340:692–696[CrossRef][Medline]
.
J Biol Chem 264:15475–15482 and Gs that alter the fidelity of receptor activation. Mol
Pharmacol 50:885–890[Abstract]
mutant in a patient with
Albright hereditary osteodystrophy uncouples cell surface receptors
from adenylyl cyclase. J Biol Chem 269:25387–25391 in which substitutions decrease
receptor-mediated activation and increase receptor affinity. Mol
Pharmacol 57:1081–10922-adrenergic inhibition of adenylyl cyclase in
platelet membranes: in situ identification with
G C-terminal antibodies. Proc Natl Acad Sci
USA 86:7809–7813s
carboxyl-terminal decapeptide. FEBS Lett 249:189–194[CrossRef][Medline]
s carboxyl-terminal
peptide prevents Gs activation by the
A2A adenosine receptor. Mol Pharmacol 58:226–236. Cell 68:911–922[CrossRef][Medline]
mutants with decreased ability to activate adenylylcyclase.
J Biol Chem 266:16226–16231 subunit gene (GNAS1) to the distal long arm of
chromosome 20 using a polymorphism detected by denaturing gradient gel
electrophoresis. Genomics 9:782–783[CrossRef][Medline]
(GNAS1), the probable candidate gene for Albright
hereditary osteodystrophy, is assigned to human chromosome
20q12–q13.2. Genomics 10:257–261[CrossRef][Medline]
subunit of the stimulatory G protein of
adenylyl cyclase (GNAS1) to 20q13.2-q13.3 in human by
in situ hybridization. Genomics 11:478–479[Medline]
splice variants on
ß2-adrenoreceptor-mediated signaling. The ß2-adrenoreceptor coupled
to the long splice variant of Gs has
properties of a constitutively active receptor. J Biol Chem 273:5109–5116 transcript. J Biol Chem 268:9879–9885s
is a new type of G protein. Nature 372:804–809[Medline]
-subunit
XLs. I. Tissue distribution and subcellular localization. J
Biol Chem 275:33622–33632 signal transduction
protein cDNA species. Nucleic Acids Res 19:4725–4729 mRNA. J Biol Chem 265:8458–8462-subunit XLs in neuroepithelial cells and
young neurons during development of the rat nervous system. Neurosci
Lett 301:119–122[CrossRef][Medline]
s to the Golgi complex region. Mol
Biol Cell 11:1421–1432-subunit XLs. II. Signal transduction properties. J
Biol Chem 275:33633–33640-subunit (Gs) knockout mice is due to
tissue-specific imprinting of the Gs gene.
Proc Natl Acad Sci USA 95:8715–8720 gene GNAS1 in the pathogenesis
of acromegaly. J Clin Invest 107:R31–R36
subunit knockout produces opposite effects on energy metabolism. J
Clin Invest 105:615–623[Medline]
subunit of Gs. Adv Second
Messenger Phosphoprotein Res 24:70–75[Medline]
gene are
associated with low levels of Gs protein in growth hormone-secreting
tumors. J Clin Endocrinol Metab 83:4386–4390 is associated with an
increased phosphodiesterase activity in human growth hormone-secreting
adenomas. J Clin Endocrinol Metab 83:1624–1628-gene in nonfunctioning pituitary tumors.
J Clin Endocrinol Metab 77:765–769[Abstract]
and Gi2 mutations in
clinically non-functioning pituitary tumours. Clin Endocrinol (Oxf) 41:815–820[Medline]
-q, G-11, G-s, or thyrotropin-releasing
hormone receptor genes. J Clin Endocrinol Metab 81:1134–1140[Abstract]
s stimulates growth and differentiation of thyroid FRTL5
cells. Oncogene 9:3647–3653[Medline]
subunit
gene in human endocrine tumors: mutation detection by polymerase chain
reaction-primer-introduced restriction analysis. Cancer 72:1386–1393[CrossRef][Medline]
and Gi2 genes in sporadic parathyroid adenomas
and insulinomas. Clin Sci 87:493–497[Medline]
s as the molecular
basis for the McCune-Albright syndrome. Arch Med Res 30:522–531[CrossRef][Medline]
in McCune-Albright syndrome causes skin pigmentation by
tyrosinase gene activation on affected melanocytes. Horm Res 52:235–240[CrossRef][Medline]
GT progressive cholestasis. J
Hepatol 32:154–158[Medline]
mutation in monostotic and polyostotic fibrous dysplasia. Am
J Pathol 150:1059–1069[Abstract]
mutation is present in
fibrous dysplasia of bone in the McCune-Albright syndrome. J Clin
Endocrinol Metab 79:750–755[Abstract]
subunit of
the stimulatory G protein of adenylyl cyclase in McCune-Albright
syndrome. Proc Natl Acad Sci USA 89:5152–5156 mutation.
Am J Hum Genet 65(Suppl):A426 (Abstract)
in adult mouse cardiomyocytes.
J Biol Chem 275:40635–40640 mutation in
intramuscular myxomas with and without fibrous dysplasia of bone.
Virchows Arch 437:133–137[CrossRef][Medline]
gene:
site-specific patterns and recurrent histological hallmarks. J
Pathol 187:249–258[CrossRef][Medline]
-subunit from a bone lesion. J Clin
Endocrinol Metab 78:803–806[Abstract]
gene. Hum Mol Genet 4:1675–1676s from patients with
fibrous dysplasia of bone. Bone 21:201–206[Medline]
-mutated skeletal progenitor cells. J Clin Invest 101:1737–1744[Medline]
promote osteoblast differentiation.
Proc 77th Meeting of The Endocrine Society, Washington, DC, 1995, p 62
s:
chronic transcriptional activation of the c-fos
proto-oncogene in endocrine cells. J Biol Chem 269:22663–22671-subunit gene in Albright hereditary
osteodystrophy detected by denaturing gradient gel electrophoresis.
Proc Natl Acad Sci USA 87:8287–8290 subunit of the stimulatory G
protein of adenylyl cyclase in Albright hereditary osteodystrophy.
J Clin Endocrinol Metab 76:1560–1568[Abstract]
gene (GNAS1) in
Albright hereditary osteodystrophy. J Clin Endocrinol Metab 84:3254–3259,25-dihydroxyvitamin D
in response to parathyroid extract in pseudohypoparathyroidism. J
Clin Invest 66:782–791
,25-Dihydroxyvitamin D3 corrects
osteomalacia in hypoparathyroidism and pseudohypoparathyroidism. Acta
Endocrinol (Copenh) 103:241–247
deficiency. J Clin
Endocrinol Metab 82:247–250 subunit of the guanine nucleotide-binding protein
Gs as the molecular basis for Albright hereditary
osteodystrophy. Proc Natl Acad Sci USA 85:617–621
subunit of the stimulatory GTP-binding protein in
pseudohypoparathyroidism type Ia. Proc Natl Acad Sci USA 84:7266–7269subunit of the stimulatory G-protein of adenylyl cyclase
in patients with Albright’s hereditary osteodystrophy. J Clin
Endocrinol Metab 71:1208–1214 gene (GNAS1) in a patient with
Albright hereditary osteodystrophy. Genomics 13:1319–1321[CrossRef][Medline]
gene (GNAS1) in Albright hereditary
osteodystrophy. Genomics 21:455–457[CrossRef][Medline]
gene mutations in
Albright’s hereditary osteodystrophy. J Med Genet 31:835–839 gene
(GNAS1) in patients with Albright hereditary osteodystrophy.
Hum Mol Genet 4:2001–2002 in patients with gain and
loss of endocrine function. Nature 371:164–167[CrossRef][Medline]
contact region of Gs impairs
receptor stimulation. J Biol Chem 271:19653–19655 gene. Hum Genet 97:73–75[Medline]
-subunit of the guanine nucleotide-binding protein
Gs in pseudo- and pseudopseudohypoparathyroidism.
J Clin Endocrinol Metab 83:935–938 in a patient with pseudohypoparathyroidism.
Mol Endocrinol 11:1718–1727 gene in a Japanese
patient with pseudohypoparathyroidism. J Endocrinol Invest 19:236–241[Medline]
gene
mutation. Am J Med Genet 77:61–67[CrossRef]
s. A cause of pseudohypoparathyroidism type
Ib. J Biol Chem 276:165–1711 as an
approximation of the receptor-bound state. J Biol Chem 273:21752–21758-subunit within different segments of the nephron. Am J Physiol
278:F507–F514
.
Am J Physiol 277:F235–F244
2-adrenergic receptors in adipose tissue of
ß3-adrenergic receptor deficient mice promotes diet-induced obesity.
J Biol Chem 275:34797–34802-subunit sequence
accelerate differentiation of fibroblasts to adipocytes. Nature 358:334–337[CrossRef][Medline]
-subunit of the stimulatory G protein and severe extraskeletal
ossification. J Bone Miner Res 15:2074–2083[CrossRef][Medline]
gene: How does this relate to hormone
resistance in Albright hereditary osteodystophy? Genomics 36:280–287[CrossRef][Medline]
knockout mice. J Biol Chem 276:19994–19998This article has been cited by other articles:
 |
|
 |
|
  L. S Kirschner, Z. Yin, G. N Jones, and E. Mahoney Mouse models of altered protein kinase A signaling Endocr. Relat. Cancer, September 1, 2009; 16(3): 773 - 793. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 F Palos-Paz, O Perez-Guerra, J Cameselle-Teijeiro, C Rueda-Chimeno, F Barreiro-Morandeira, J Lado-Abeal, the Galician Group for the Study of Toxic Multinod, D Araujo Vilar, R Argueso, O Barca, et al. Prevalence of mutations in TSHR, GNAS, PRKAR1A and RAS genes in a large series of toxic thyroid adenomas from Galicia, an iodine-deficient area in NW Spain Eur. J. Endocrinol., November 1, 2008; 159(5): 623 - 631. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A.-S. Balavoine, M. Ladsous, F.-L. Velayoudom, V. Vlaeminck, C. Cardot-Bauters, M. d'Herbomez, and J.-L. Wemeau Hypothyroidism in patients with pseudohypoparathyroidism type Ia: clinical evidence of resistance to TSH and TRH Eur. J. Endocrinol., October 1, 2008; 159(4): 431 - 437. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. Krechowec and A. Plagge Physiological Dysfunctions Associated with Mutations of the Imprinted Gnas Locus Physiology, August 1, 2008; 23(4): 221 - 229. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 V. Mariot, S. Maupetit-Mehouas, C. Sinding, M.-L. Kottler, and A. Linglart A Maternal Epimutation of GNAS Leads to Albright Osteodystrophy and Parathyroid Hormone Resistance J. Clin. Endocrinol. Metab., March 1, 2008; 93(3): 661 - 665. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Plagge, G. Kelsey, and E. L Germain-Lee Physiological functions of the imprinted Gnas locus and its protein variants G{alpha}s and XL{alpha}s in human and mouse J. Endocrinol., February 1, 2008; 196(2): 193 - 214. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 Z. Yin, L. Williams-Simons, A. F. Parlow, S. Asa, and L. S. Kirschner Pituitary-Specific Knockout of the Carney Complex Gene Prkar1a Leads to Pituitary Tumorigenesis Mol. Endocrinol., February 1, 2008; 22(2): 380 - 387. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 N. Ogata, H. Kawaguchi, U.-i. Chung, S. I. Roth, and G. V. Segre Continuous Activation of G{alpha}q in Osteoblasts Results in Osteopenia through Impaired Osteoblast Differentiation J. Biol. Chem., December 7, 2007; 282(49): 35757 - 35764. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 T. Xie, M. Chen, Q.-H. Zhang, Z. Ma, and L. S. Weinstein cell-specific deficiency of the stimulatory G protein {alpha}-subunit Gs{alpha} leads to reduced cell mass and insulin-deficient diabetes PNAS, December 4, 2007; 104(49): 19601 - 19606. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. Mantovani, S. Bondioni, A. Linglart, M. Maghnie, M. Cisternino, S. Corbetta, A. G. Lania, P. Beck-Peccoz, and A. Spada Genetic Analysis and Evaluation of Resistance to Thyrotropin and Growth Hormone-Releasing Hormone in Pseudohypoparathyroidism Type Ib J. Clin. Endocrinol. Metab., September 1, 2007; 92(9): 3738 - 3742. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. P. de Nanclares, E. Fernandez-Rebollo, I. Santin, B. Garcia-Cuartero, S. Gaztambide, E. Menendez, M. J. Morales, M. Pombo, J. R. Bilbao, F. Barros, et al. Epigenetic Defects of GNAS in Patients with Pseudohypoparathyroidism and Mild Features of Albright's Hereditary Osteodystrophy J. Clin. Endocrinol. Metab., June 1, 2007; 92(6): 2370 - 2373. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 L. F. Frohlich, M. Bastepe, D. Ozturk, H. Abu-Zahra, and H. Juppner Lack of Gnas Epigenetic Changes and Pseudohypoparathyroidism Type Ib in Mice with Targeted Disruption of Syntaxin-16 Endocrinology, June 1, 2007; 148(6): 2925 - 2935. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. N. Long, S. McGuire, M. A. Levine, L. S. Weinstein, and E. L. Germain-Lee Body Mass Index Differences in Pseudohypoparathyroidism Type 1a Versus Pseudopseudohypoparathyroidism May Implicate Paternal Imprinting of G{alpha}s in the Development of Human Obesity J. Clin. Endocrinol. Metab., March 1, 2007; 92(3): 1073 - 1079. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
F. Galland, P. Kamenicky, H. Affres, Y. Reznik, D. Pontvert, Y. Le Bouc, J. Young, and P. Chanson McCune-Albright Syndrome and Acromegaly: Effects of Hypothalamopituitary Radiotherapy and/or Pegvisomant in Somatostatin Analog-Resistant Patients J. Clin. Endocrinol. Metab., December 1, 2006; 91(12): 4957 - 4961. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Hahn, U. H Frey, W. Siffert, S. Tan, K. Mann, and O. E Janssen The CC genotype of the GNAS T393C polymorphism is associated with obesity and insulin resistance in women with polycystic ovary syndrome. Eur. J. Endocrinol., November 1, 2006; 155(5): 763 - 770. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. Xie, A. Plagge, O. Gavrilova, S. Pack, W. Jou, E. W. Lai, M. Frontera, G. Kelsey, and L. S. Weinstein The Alternative Stimulatory G Protein {alpha}-Subunit XL{alpha}s Is a Critical Regulator of Energy and Glucose Metabolism and Sympathetic Nerve Activity in Adult Mice J. Biol. Chem., July 14, 2006; 281(28): 18989 - 18999. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 E. W. Hillhouse and D. K. Grammatopoulos The Molecular Mechanisms Underlying the Regulation of the Biological Activity of Corticotropin-Releasing Hormone Receptors: Implications for Physiology and Pathophysiology Endocr. Rev., May 1, 2006; 27(3): 260 - 286. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Linglart, M. J. Mahon, M. A. Kerachian, D. M. Berlach, G. N. Hendy, H. Juppner, and M. Bastepe Coding GNAS Mutations Leading to Hormone Resistance Impair in Vitro Agonist- and Cholera Toxin-Induced Adenosine Cyclic 3',5'-Monophosphate Formation Mediated by Human XL{alpha}s Endocrinology, May 1, 2006; 147(5): 2253 - 2262. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. Bastepe and H. Juppner Pseudohypoparathyroidism, Gs{alpha}, and the GNAS Locus IBMS BoneKEy, December 1, 2005; 2(12): 20 - 32. [Full Text] [PDF]
|
|
|
|
 |
|
 K. S. Nadella and L. S. Kirschner Disruption of Protein Kinase A Regulation Causes Immortalization and Dysregulation of D-Type Cyclins Cancer Res., November 15, 2005; 65(22): 10307 - 10315. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. L. Germain-Lee, W. Schwindinger, J. L. Crane, R. Zewdu, L. S. Zweifel, G. Wand, D. L. Huso, M. Saji, M. D. Ringel, and M. A. Levine A Mouse Model of Albright Hereditary Osteodystrophy Generated by Targeted Disruption of Exon 1 of the Gnas Gene Endocrinology, November 1, 2005; 146(11): 4697 - 4709. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 N. Wettschureck and S. Offermanns Mammalian G Proteins and Their Cell Type Specific Functions Physiol Rev, October 1, 2005; 85(4): 1159 - 1204. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. J. Marx and W. F. Simonds Hereditary Hormone Excess: Genes, Molecular Pathways, and Syndromes Endocr. Rev., August 1, 2005; 26(5): 615 - 661. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Sakamoto, M. Chen, T. Nakamura, T. Xie, G. Karsenty, and L. S. Weinstein Deficiency of the G-protein {alpha}-Subunit Gs{alpha} in Osteoblasts Leads to Differential Effects on Trabecular and Cortical Bone J. Biol. Chem., June 3, 2005; 280(22): 21369 - 21375. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Chen, O. Gavrilova, J. Liu, T. Xie, C. Deng, A. T. Nguyen, L. M. Nackers, J. Lorenzo, L. Shen, and L. S. Weinstein Alternative Gnas gene products have opposite effects on glucose and lipid metabolism PNAS, May 17, 2005; 102(20): 7386 - 7391. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. Liu, M. Chen, C. Deng, D. Bourc'his, J. G. Nealon, B. Erlichman, T. H. Bestor, and L. S. Weinstein From the Cover: Identification of the control region for tissue-specific imprinting of the stimulatory G protein {alpha}-subunit PNAS, April 12, 2005; 102(15): 5513 - 5518. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 F. H. Mahmud, A. Linglart, M. Bastepe, H. Juppner, and A. N. Lteif Molecular Diagnosis of Pseudohypoparathyroidism Type Ib in a Family With Presumed Paroxysmal Dyskinesia Pediatrics, February 1, 2005; 115(2): e242 - e244. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Liu, J. G. Nealon, and L. S. Weinstein Distinct patterns of abnormal GNAS imprinting in familial and sporadic pseudohypoparathyroidism type IB Hum. Mol. Genet., January 1, 2005; 14(1): 95 - 102. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. Laspa, M. Bastepe, H. Juppner, and A. Tsatsoulis Phenotypic and Molecular Genetic Aspects of Pseudohypoparathyroidism Type Ib in a Greek Kindred: Evidence for Enhanced Uric Acid Excretion Due to Parathyroid Hormone Resistance J. Clin. Endocrinol. Metab., December 1, 2004; 89(12): 5942 - 5947. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. Mantovani, S. Bondioni, M. Locatelli, C. Pedroni, A. G. Lania, E. Ferrante, M. Filopanti, P. Beck-Peccoz, and A. Spada Biallelic Expression of the Gs{alpha} Gene in Human Bone and Adipose Tissue J. Clin. Endocrinol. Metab., December 1, 2004; 89(12): 6316 - 6319. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. S. Weinstein, J. Liu, A. Sakamoto, T. Xie, and M. Chen Minireview: GNAS: Normal and Abnormal Functions Endocrinology, December 1, 2004; 145(12): 5459 - 5464. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Bastepe, L. S. Weinstein, N. Ogata, H. Kawaguchi, H. Juppner, H. M. Kronenberg, and U.-i. Chung Stimulatory G protein directly regulates hypertrophic differentiation of growth plate cartilage in vivo PNAS, October 12, 2004; 101(41): 14794 - 14799. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Chen, M. Haluzik, N. J. Wolf, J. Lorenzo, K. R. Dietz, M. L. Reitman, and L. S. Weinstein Increased Insulin Sensitivity in Paternal Gnas Knockout Mice Is Associated with Increased Lipid Clearance Endocrinology, September 1, 2004; 145(9): 4094 - 4102. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. Abramowitz, D. Grenet, M. Birnbaumer, H. N. Torres, and L. Birnbaumer XL{alpha}s, the extra-long form of the {alpha}-subunit of the Gs G protein, is significantly longer than suspected, and so is its companion Alex PNAS, June 1, 2004; 101(22): 8366 - 8371. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Sakamoto, J. Liu, A. Greene, M. Chen, and L. S. Weinstein Tissue-specific imprinting of the G protein Gs{alpha} is associated with tissue-specific differences in histone methylation Hum. Mol. Genet., April 15, 2004; 13(8): 819 - 828. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J.-C. Carel, N. Lahlou, M. Roger, and J. L. Chaussain Precocious puberty and statural growth Hum. Reprod. Update, March 1, 2004; 10(2): 135 - 147. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Bastepe and H. Juppner Pseudohypoparathyroidism and Mechanisms of Resistance toward Multiple Hormones: Molecular Evidence to Clinical Presentation J. Clin. Endocrinol. Metab., September 1, 2003; 88(9): 4055 - 4058. [Full Text] [PDF]
|
|
|
|
|
|
E. L. Germain-Lee, J. Groman, J. L. Crane, S. M. Jan de Beur, and M. A. Levine Growth Hormone Deficiency in Pseudohypoparathyroidism Type 1a: Another Manifestation of Multihormone Resistance J. Clin. Endocrinol. Metab., September 1, 2003; 88(9): 4059 - 4069. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. Mantovani, M. Maghnie, G. Weber, E. De Menis, V. Brunelli, M. Cappa, P. Loli, P. Beck-Peccoz, and A. Spada Growth Hormone-Releasing Hormone Resistance in Pseudohypoparathyroidism Type Ia: New Evidence for Imprinting of the Gs{alpha} Gene J. Clin. Endocrinol. Metab., September 1, 2003; 88(9): 4070 - 4074. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. Liu, B. Erlichman, and L. S. Weinstein The Stimulatory G Protein {alpha}-Subunit Gs{alpha} Is Imprinted in Human Thyroid Glands: Implications for Thyroid Function in Pseudohypoparathyroidism Types 1A and 1B J. Clin. Endocrinol. Metab., September 1, 2003; 88(9): 4336 - 4341. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C. Coombes, P. Arnaud, E. Gordon, W. Dean, E. A. Coar, C. M. Williamson, R. Feil, J. Peters, and G. Kelsey Epigenetic Properties and Identification of an Imprint Mark in the Nesp-Gnasxl Domain of the Mouse Gnas Imprinted Locus Mol. Cell. Biol., August 15, 2003; 23(16): 5475 - 5488. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Demura, Y. Takeda, T. Yoneda, K. Furukawa, A. Tachi, and H. Mabuchi Completely Skewed X-Inactivation in a Mentally Retarded Young Female with Pseudohypoparathyroidism Type IB and Juvenile Renin-Dependent Hypertension J. Clin. Endocrinol. Metab., July 1, 2003; 88(7): 3043 - 3049. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 F Sandrini, L S Kirschner, T Bei, C Farmakidis, J Yasufuku-Takano, K Takano, T R Prezant, S J Marx, W E Farrell, R N Clayton, et al. PRKAR1A, one of the Carney complex genes, and its locus (17q22-24) are rarely altered in pituitary tumours outside the Carney complex J. Med. Genet., December 1, 2002; 39(12): e78 - 78. [Full Text] [PDF]
|
|
|
|
|
|
K. Freson, C. Thys, C. Wittevrongel, W. Proesmans, M. F. Hoylaerts, J. Vermylen, and C. Van Geet Pseudohypoparathyroidism type Ib with disturbed imprinting in the GNAS1 cluster and Gs{alpha} deficiency in platelets Hum. Mol. Genet., October 15, 2002; 11(22): 2741 - 2750. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Bastepe, Y. Gunes, B. Perez-Villamil, J. Hunzelman, L. S. Weinstein, and H. Juppner Receptor-Mediated Adenylyl Cyclase Activation Through XL{alpha}s, the Extra-Large Variant of the Stimulatory G Protein {alpha}-Subunit Mol. Endocrinol., August 1, 2002; 16(8): 1912 - 1919. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 S. F. Steinberg Focus on "Targeted expression of activated Q227L Galpha s in vivo" Am J Physiol Cell Physiol, August 1, 2002; 283(2): C383 - C385. [Full Text] [PDF]
|
|
|
|
|
|
P. Kopp Perspective: Genetic Defects in the Etiology of Congenital Hypothyroidism Endocrinology, June 1, 2002; 143(6): 2019 - 2024. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 H. Juppner The Genetic Basis of Progressive Osseous Heteroplasia N. Engl. J. Med., January 10, 2002; 346(2): 128 - 130. [Full Text] [PDF]
|
|
|
|
|
|
L. S. Weinstein The Stimulatory G Protein {alpha}-Subunit Gene: Mutations and Imprinting Lead to Complex Phenotypes J. Clin. Endocrinol. Metab., October 1, 2001; 86(10): 4622 - 4626. [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
| Endocrinology | Endocrine Reviews | J. Clin. End. & Metab. |
| Molecular Endocrinology | Recent Prog. Horm. Res. | All Endocrine Journals |
