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Journal of Clinical Endocrinology & Metabolism, doi:10.1210/jc.2006-2122
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The Journal of Clinical Endocrinology & Metabolism Vol. 92, No. 5 1764-1768
Copyright © 2007 by The Endocrine Society

A Disruptive Mutation in Exon 3 of the GNAS Gene with Albright Hereditary Osteodystrophy, Normocalcemic Pseudohypoparathyroidism, and Selective Long Transcript Variant Gs{alpha}-L Deficiency

Susanne Thiele, Ralf Werner, Wiebke Ahrens, Ute Hoppe, Christine Marschke, Pia Staedt and Olaf Hiort

Department of Pediatrics and Adolescent Medicine, University of Luebeck, 23538 Luebeck, Germany

Address all correspondence and requests for reprints to: Olaf Hiort, M.D., Division of Pediatric Endocrinology and Diabetes, Department of Pediatrics and Adolescent Medicine, University of Luebeck, Ratzeburger Allee 160, 23538 Luebeck, Germany. E-mail: hiort{at}paedia.ukl.mu-luebeck.de.


   Abstract
Top
 Abstract
Introduction
 Patients and Methods
 Results
 Discussion
 References
 
Objective: The GNAS gene encodes the {alpha}-subunit of stimulatory G proteins, which play a crucial role in intracellular signal transduction of peptide and neurotransmitter receptors. In addition to transcript variants that differ in their first exon due to different promoters, there are two long (Gs{alpha}-L) and two short (Gs{alpha}-S) splice variants, created by alternative splicing. Heterozygous inactivating maternally inherited mutations of GNAS lead to a phenotype in which Albright hereditary osteodystrophy is associated with pseudohypoparathyroidism type Ia.

Methods and Results: The GNAS gene of a 10-yr-old girl with brachymetacarpia, mental retardation, normocalcemic pseudohypoparathyroidism, and hypothyroidism was investigated. We found a heterozygous insertion of an adenosine in exon 3 altering codon 85 and leading to a frame shift inducing a stop codon in exon 4. Molecular studies of cDNA from blood RNA demonstrated normal, biallelic expression of Gs{alpha}-S transcripts, whereas expression of Gs{alpha}-L transcripts from the maternal allele was reduced. Immunoblot analysis revealed a reduced Gs{alpha}-L protein level to about 50%, whereas the protein level of Gs{alpha}-S was unaltered. Furthermore, the Gs{alpha} protein activity in erythrocyte membranes was diminished to about 75% of normal. Both the reduced activity and the mutation were also found in the mother and the affected younger brother.

Conclusion: This report demonstrates the first evidence for a pathogenic mutation in exon 3 of the GNAS gene. The mutation is associated with a phenotype of Albright hereditary osteodystrophy and pseudohypoparathyroidism type Ia due to selective deficiency of Gs{alpha}-L and a partial reduction of Gs{alpha} activity.


   Introduction
Top
 Abstract
 Introduction
Patients and Methods
 Results
 Discussion
 References
 
GUANINE NUCLEOTIDE BINDING proteins are cytoplasmatic membrane-associated heterotrimers, composed of an {alpha}-, ß-, and {gamma}-subunit and are crucial in coupling extracellular activated membrane receptors to intracellular second messenger enzyme systems. In the inactive state, GDP is bound to the {alpha}-subunit Gs{alpha}. When a ligand interacts with a G protein-coupled receptor (GPCR), GDP-GTP exchange on Gs{alpha} is promoted, resulting in activation and dissociation from the ß{gamma}-subunits. GDP release is assumed to be the rate-limiting step of the G protein cycle (1). Then GTP-Gs{alpha} mediates signal transduction of hormones, neurotransmitters, or glycoproteins to cAMP generation via the stimulation of adenylate cyclase. The turn-off mechanism is performed by an intrinsic GTPase that hydrolyzes bound GTP to GDP (1, 2).

Gs{alpha} is encoded by the GNAS gene, located on chromosome 20q13.11 and consists of 13 exons and 12 introns (3). Moreover, several different transcript variants from GNAS have been described (3, 4, 5, 6, 7, 8), and GNAS has been termed one of the most complex gene loci in the human genome (2). The variants are transcribed from different promoter regions [e.g. Gs{alpha}, neuroendocrine secretory protein 55 antisense (Nespas), extra large Gs{alpha} (XL{alpha}s), and exon 1a] (5, 6, 7, 8) and are alternatively spliced. By different splicing of exon 3 and/or use of two 5'splice sites of exon 4, two long (Gs{alpha}-L) and two short (Gs{alpha}-S) transcript variants are created, which contain alternatively exon 3 and/or a CAG sequence, respectively (3, 4, 9). Each of these variants represents the transcript for functioning proteins and is capable of stimulating both adenylate cyclase (10, 11) and calcium channels (11). Regarding exon 3, the same splice variants are described for XL{alpha}s (12), Nesp55 (13), and exon 1a (14, 15). However, only XL{alpha}s has a long and a short protein variant because the exon 3 coding sequence is not part of a protein coding sequence in Nesp55 and exon 1a.

On the basis of the versatility of signal transduction pathways in many different tissues that are controlled by Gs{alpha} (16), inactivating mutations of GNAS are associated with a heterogeneity of phenotypic signs. Affected tissues include bone, skin, and adipose tissue with shortening of the fourth and fifth metacarpals and metatarsals, a round face, sc calcifications, short stature, and obesity (15, 17). Additionally, mental retardation may be a sign of altered signal pathways in the brain (18). These features characterize Albright hereditary osteodystrophy (AHO), first described by Albright in 1942 (19).

GNAS is biallelically expressed in most tissues studied (15, 17); however, in some tissues GNAS is imprinted with silencing of the paternal allele (20, 21) leading to a parental-of-origin effect. In case of maternally inherited mutation, AHO is associated with end-organ resistance to the Gs{alpha}-mediated action of different hormones, primarily PTH, TSH, gonadotropin, and GHRH. AHO with endocrinopathy is then termed pseudohypoparathyroidism type Ia (PHP Ia). In contrast, AHO due to paternally inherited mutation transmission lacks biochemical evidence of hormone resistance and is designated as pseudopseudohypoparathyroidism (2, 22).

In recent years more than 80 different heterozygous inactivating mutations in the GNAS gene have been identified, which are distributed throughout the gene (23, 24, 25, 26) (www.hgmd.org). So far, no mutation has been described in exon 3, which would be predicted to selectively affect the Gs{alpha}-L isoform. This report documents the clinical, biochemical, and molecular features of a patient with a mutation in exon 3 of the GNAS gene with phenotypic signs of AHO and PHP Ia.


   Patients and Methods
Top
 Abstract
 Introduction
 Patients and Methods
Results
 Discussion
 References
 
Patients

The female patient was born at term (42 wk gestational age) after an uneventful pregnancy as the second child of unrelated Caucasian parents. Her weight at birth was 4400 g (>90th centile) and her length 57 cm (>90th centile). The neonatal period was uncomplicated and except for a mild delay of psychomotor and speech development, no further atypical signs were observed in early childhood. At the age of 5 yr, an endocrine consultation at an outside institution was carried out because of observed weight gain. At laboratory investigations, slight primary hypothyroidism with an elevated TSH to 18.9 mU/liter (reference range 0.85–6.5 mU/liter) and low free tetraiodothyronine (fT4) of 0.7 pmol/liter (reference range 12.1–22.0 pmol/liter) was found. Also, PTH was elevated to 109.3 pg/ml (reference range up to 70 pg/ml) with normal serum calcium values and normal 25-(OH) vitamin D3 levels. The diagnosis of normocalcemic pseudohypoparathyroidism and overt hypothyroidism was made and treatment with levothyroxine was initiated. Because she had not shown clinical and biochemical signs of hypocalcemia, she was not treated with calcitriol. Because of suspicion of AHO, she was referred to our institution for further evaluation at the age of 101/2 yr. At this time she showed features of AHO with a round face and brachymetacarpia of the fourth and fifth digits, but she did not have any sc calcifications. Her weight was 37.8 kg (25th centile) and her length 137.5 cm (25th centile); thus, there was a normal body mass index at this time (20 kg/m2, < 90th centile). By radiography, her bone age was accelerated to 13 yr 6 months (Greulich and Pyle). She was receiving 50 µg levothyroxine per day.

Her 9-month-old brother was also presented. He was born after an uneventful pregnancy as the family’s third child at 35 wk gestation. His birth weight was 3000 g (>90th percentile), his birth length 48 cm (>90th percentile). He showed slight psychomotor retardation and one small sc calcification on his left cheek.

Both her parents and her older brother did not show any visible signs of AHO. The parents gave informed consent to further investigations in all family members. Studies were approved by the ethical committee of the University of Luebeck as part of the funded project on AHO (see Acknowledgments).

Laboratory investigations

Laboratory tests of the girl revealed a normocalcemic pseudohypoparathyroidism characterized by a slightly elevated PTH, a serum calcium in the mid-normal range, and a slightly decreased serum phosphate (Table 1Go). The IGF-I level as a peripheral marker of human GH secretion was decreased. The hypothyroidism was sufficiently treated because TSH, fT4, and free triiodothyronine (fT3) levels were now in the normal range (Table 1Go).


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  		TABLE 1. Laboratory investigations of our patient at the age of 10 yr and her younger brother at the age of 9 months

 
The affected brother demonstrated hyperthyreotropinemia with increased TSH but normal fT4 and fT3 levels. IGF-I was also just below the normal range. PTH, calcium, and phosphate were in the middle of the reference range; therefore, there was no evidence for a disturbance in calcium-phosphate metabolism at this time (Table 1Go). The mother had normal values for calcium, phosphate, and PTH in serum.

Gs{alpha} protein activity

The activity of Gs{alpha} protein was investigated in erythrocyte membranes from EDTA blood samples as described earlier (22) based on the method described by Levine et al. (27) and adapted by Marguet et al. (28). After solubilization and activation, we measured the activation of the Gs{alpha} protein via the generation of cAMP using adenylate cyclase from turkey red cell membranes in presence of ATP by RIA (RIA-Kit RE 11021; IBL, Hamburg, Germany). Results obtained in triplicate were expressed as percent of the mean of healthy controls. The Gs{alpha} activity of the girl’s sample was measured in triplicate in two independent blood samples.

Molecular genetic analysis

Genomic DNA derived from peripheral leukocytes was isolated by standard procedures (Qiaquick DNA kit; QIAGEN, Hilden, Germany). Exon 1–13 of the GNAS gene including intron/exon boundaries were amplified in 13 fragments by PCR using the oligonucleotide primers reported by Ahrens et al. (24). Mutation screening was carried out by a nonisotopic single-strand conformation polymorphism (SSCP) analysis on 10% polyacrylamide gel as described earlier (24). Samples of the patient, her mother, and her younger brother showing an aberrant migration pattern of the exon 3 PCR product, compared with control persons, were reamplified from genomic DNA and sequenced. For FokI-polymorphism analysis, we also sequenced exon 5 in samples from the parents and the affected children. Sequence analysis of DNA was performed with CY5-labeled primers in sense and antisense direction analyzed with an automatic sequencer (ALF Express II; Amersham Pharmacia, Freiburg, Germany) using SequiTherm EXEL II Long-Read DNA Sequencing Kit Alf (Biozym, Hess, Oldendorf, Germany), according to the directions provided by the manufacturer.

RT-PCR of Gs{alpha} mRNA

Total RNA from blood samples of our patient was collected in a PAXgene blood RNA tube and isolated using the PAXgene blood RNA kit (QIAGEN). Reverse transcription was achieved by using 1 µg of total RNA and 20 pmol of random primers using Superscript II RT (Invitrogen, Mannheim, Germany) according to the instructions of the manufacturer. The cDNAs were amplified by PCR using the exon 1-specific sense primer 5'-CCATGGGCTGCCTCGGGAACA-3' and the exon 6-specific antisense primer 5'-CCTTGGCATGCTCATAGAATTC-3'. After an initial denaturation at 94 C for 5 min, 34 cycles of PCR amplification were performed with each cycle consisting of 1.30 min for annealing at 62 C, 2.0 min at 72 C for primer extension, and 1.15 min at 94 C for denaturation.

Allele-specific expression analysis

The cDNAs of our index patient and her parents were amplified using the exon 1-specific sense and the exon 6-specific antisense primer. Subsequently the amplicons were subjected to FokI digestion and separated on a 2% agarose gel. Amplified genomic DNA containing exon 3 served as a positive hybridization control, whereas an amplified cDNA containing exon 1–6 of the Gs{alpha}-S transcript (without exon 3) served as a negative control. Amplified DNA fragments were then transferred to a nylon membrane (Hybond NX; Amersham Pharmacia, Birmingham, UK) overnight using the TURBO Blotter device (Whatman, Dassel, Germany). Hybridization was carried out with a digoxigenin (DIG)-labeled exon 3-specific oligonucleotide probe (5'-DIG-GGGCGGCGAAGAGGACCCGCAGGCTGCAAGGAGCAACAGCGATGG-3'; Metabion, Martinsried, Germany) at 47 C for 2.5 h. Membranes were washed twice with 2x saline sodium citrate and 0.1% sodium dodecyl sulfate at room temperature for 5 min and twice with 0.1x saline sodium citrate and 0.1% sodium dodecyl sulfate at 68 C for 15 min. Bands were visualized using anti-DIG AP fab fragments according to the instructions provided by the manufacturer (Roche, Mannheim, Germany).

Immunoblot analysis

Protein samples of our patient and three adult controls were solubilized from erythrocyte membranes as described above in the Gs{alpha} activity section. Fifteen micrograms of protein were loaded onto a discontinuous 12% polyacrylamide gel. Electrophoresis was carried out at 100 V for 3 h in a Mini-Protean 3 chamber (Bio-Rad, Munich, Germany). Proteins were transferred to nitrocellulose membranes (BA85; Schleicher & Schüll, Dassel, Germany) for 2.5 h in a Mini Trans-Blot cell (Bio-Rad) using a blotting buffer containing 16.5 mM Tris, 150 mM glycine, and 20% (vol/vol) methanol. Nonspecific binding sites were blocked by immersing the membranes in 5% dried skimmed milk (Becton Dickinson, Franklin Lakes, NJ) made up in PBS/Tween 20 buffer [containing 137 mM NaCl, 2.7 mM KCl, 10 mM Na2HPO4, 2 mM KH2PO4/0.1% Tween 20 (pH 7.4)] overnight at 4 C. Then the blocked membranes were incubated with a polyclonal rabbit anti-Gs{alpha} antibody generated against the synthetic C-terminal Gs{alpha} decapeptid RMHLRQYELL (Upstate Biotechnology, Lake Placid, NY) diluted 1:1000 in 5% blocking milk for 1 h at room temperature with gentle agitation. The membrane was washed four times for 15 min in PBS Tween 20 buffer and subsequently incubated with an antirabbit IgG peroxidase conjugate diluted 1:2000 (Sigma, Taufkirchen, Germany) for 1 h. After washing the membrane four times as before, protein bands were visualized using the Western Lightning Chemiluminescence Reagent Plus substrate (PerkinElmer, Boston, MA) and the Chemidoc EQ detection system (Bio-Rad). The 28-kDa membrane protein aquaporin was detected with mouse antiaquaporin 1 diluted 1:1000 (MCA2099; Serotec, Düsseldorf, Germany) and antimouse peroxidase antibodies diluted 1:2000 (Amersham Biosciences, Freiburg, Germany).


   Results
Top
 Abstract
 Introduction
 Patients and Methods
 Results
Discussion
 References
 
We measured the protein activity of Gs{alpha} from erythrocyte membranes of the patient in two different samples in triplicate. The first average result was 77.3% (73.3–81.8%), and the second was 71.4% (68–71.6%), compared with healthy controls, confirming the presumptive diagnosis of PHP Ia. The samples of her mother and her younger brother showed also a reduced Gs{alpha} activity to 71.9% (62.1–80.7%) and 67.5% (65.5–70.3%), respectively.

By SSCP analysis we found an aberrant migration pattern in the amplicon of exon 3 (Fig. 1AGo) and sequencing revealed an insertion of an adenosine in exon 3 at codon position 85 (Fig. 1BGo), giving rise to a stop codon in codon 87 in exon 4. We also documented the presence of the same mutation in the samples of the mother and younger brother of our index patient (data not shown). The PCR products of the father and healthy older brother did not show any aberrant migration pattern on SSCP analysis (see Fig. 1AGo).


Figure 1
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  		FIG. 1. A, Single-strand conformation analysis of exon 3 of GNAS revealed two bands instead of one in samples of our patient (1 ), her mother (2 ), and her younger brother (3 ), indicating a heterozygous status. Samples of her father (4 ), her older brother (5 ), and a normal control (6 ) showed a single band. B, Sequence analysis of the patients exon 3 revealed a heterozygous insertion of an adenosine (red arrow) at codon 85, resulting in a frame shift. Mutated and normal allele are visualized by double peaks from the position of the insertion. The upper letters in bold denote the wild-type sequence, the lower letters the aberrant sequence. Capital letters represent the coding sequence; lowercase letters represent intron 3 sequences.

 
Sequencing of the mother’s and father’s alleles from genomic DNA revealed the father being heterozygous (FokI+/FokI–) and the mother being homozygous (FokI+/FokI+) for the known silent FokI polymorphism in exon 5 with a single nucleotide replacement of cytosine instead of thymine in codon 131 (ATT-ATC) (27). By analysis of the short splice variants from the patient’s blood derived mRNA, we detected the unchanged Gs{alpha}-S variants and the FokI polymorphism in a heterozygous fashion (data not shown). This pattern implies biallelic expression of Gs{alpha}-S and proves the existence of normal Gs{alpha}-S transcripts from not only the paternal but also the maternal allele.

To investigate the effects of the nonsense mutation on Gs{alpha}-L mRNA accumulation, we performed hybridization analysis of amplified Gs{alpha}-L cDNA fragments containing exons 1–6 using a DIG-labeled exon 3 probe in samples of our patient and her parents. Restriction analysis of the cDNA samples of her parents confirmed heterozygosity in the father and homozygosity in the mother for the common FokI polymorphism. We also demonstrated the existence of Gs{alpha}-L with and without the FokI polymorphism and therefore biallelic expression in our patient and her father. However, in the patient only minute amounts of the maternal Gs{alpha}-L transcript (FokI+) were seen, whereas the paternally derived transcript (FokI–) was of normal amount (Fig. 2Go). This implies the suspected nonsense-mediated mRNA decay of the mutation bearing maternal transcript of Gs{alpha}-L.


Figure 2
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  		FIG. 2. Allele-specific expression of Gs{alpha}-L. Exon 1–6 of Gs{alpha} cDNAs were amplified and loaded onto a 2% agarose gel as undigested amplicons (456 bp) (a) or after FokI digestion (b). The Gs{alpha}-L-specific amplicon was detected by hybridization with an exon 3-specific probe. Lane 1, The mother (FokI+/FokI+); lane 2, the father (FokI–/FokI+); lane 3, the propositus (FokI+/FokI–); lane 4, Gs{alpha}-S amplicon containing exons 1, 2, 4, 5, and 6 (negative hybridization control); and lane 5, amplicon from genomic DNA containing exon 3 (positive hybridization control). Hybridization and FokI enzyme restriction analysis demonstrated the existence of Gs{alpha}-L mRNA of the paternal allele and the strong reduction of the maternally derived mutation bearing transcript in our index patient.

 
Immunoblot analysis from erythrocyte membranes using an antibody specific for the C-terminal end of Gs{alpha} revealed two specific bands for Gs{alpha}-S and Gs{alpha}-L. Samples of the three normal controls showed very similar results of the calculated ratio from the long and short variants, compared with aquaporin as an internal standard. As expected, the sample of our patient revealed an approximately 50% diminished protein concentration of Gs{alpha}-L (ratio Gs{alpha}-L/aquaporin 0.13), compared with three normal controls (ratios Gs{alpha}-L to aquaporin 0.26; 0.22; 0.2; mean 0.23), whereas the protein level of Gs{alpha}-S was unaltered (Fig. 3Go).


Figure 3
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  		FIG. 3. Comparison of protein levels of Gs{alpha}-L and Gs{alpha}-S by immunoblot analysis. Lane 1, patient; lanes 2–4, normal controls. Whereas the protein level of the short variant Gs{alpha}-S was similar to that of normal controls, the concentration of the long variant Gs{alpha}-L was diminished to about 50% in the patient sample. Aquaporin was used as a loading control (lower panel).

 

   Discussion
Top
 Abstract
 Introduction
 Patients and Methods
 Results
 Discussion
References
 
Our patient demonstrates the clinical, genetic, and biochemical consequences of a nonsense mutation affecting exon 3 of the GNAS gene. In contrast to all previously reported GNAS mutations in exons 1–2 and 4–13, the described mutation in exon 3 leads to a deficiency limited to the long transcript variants Gs{alpha}-L, whereas amount and function of the short variants Gs{alpha}-S are seemingly not affected. We demonstrated the existence of the normal short splice variant mRNAs transcribed from both alleles in the patient by comparing the pattern of the well-documented FokI polymorphism (29). The Gs{alpha}-L transcripts from the maternal allele were strongly reduced, probably due to a nonsense mediated decay (30). Furthermore, the level of Gs{alpha}-L protein was reduced to about 50%, compared with controls, in contrast to normal Gs{alpha}-S, confirming the selective decrease of Gs{alpha}-L. This may explain the partially reduced Gs{alpha} activity found in the patient as well as in the affected mother and brother to about 70–75% of normal.

There are controversial discussions in the literature whether the Gs{alpha} variants may have evolved to perform different regulatory functions or whether they are normal variants interacting exactly in the same way (31). Therefore, from our study we cannot predict the precise mechanism causing the phenotype of AHO and PHP Ia by the mutation affecting only the long transcript variants.

In vitro reconstitution assays using bacterially expressed {alpha}-subunits, Gs{alpha}-L, and Gs{alpha}-S showed only subtle differences in their biochemical features (10, 11, 32). Both are capable to activate the adenylate cyclase and calcium channels (11) and have thus been regarded to be functionally nearly identical. Gs{alpha}-L and Gs{alpha}-S have also been biochemically nearly indistinguishable in a mammalian cell line that lacks endogenous Gs{alpha} (32). Gs{alpha} is transcribed biallelic in many tissues affected by AHO such as adipose tissue and bone (15). If all Gs{alpha} protein variants interact similarly, the development of AHO in our case could be explained by the quantitative loss of one fourth of the entire activity of Gs{alpha} in target tissues and/or signal pathways. Moreover, this model could be supported by the level of expression of the endocrinopathies and clinical signs of AHO in our patient because she demonstrates only normocalcemic pseudohypoparathyroidism, and at this time there is no evidence for obesity. This may be an indication that the mutation can be partly compensated by a normal function of the Gs{alpha}-S from the maternal allele. Yet also mutations in the other exons have been described with this mild phenotype, and there is only a minor genotype-phenotype correlation in AHO and PHP Ia. Another mechanism explaining the phenotype in our patient would be that tissues may express different ratios of Gs{alpha}-L and Gs{alpha}-S and that the phenotypic signs could be caused by a reduction in Gs{alpha}-L in tissues expressing predominantly this isoform. However, little is known about the relative distribution of both variants in humans in AHO-affected tissues (31).

Alternative pre-mRNA splicing is an important mechanism for generating protein diversity and may explain in part how mammalian complexity arises from a surprisingly small complement of genes (33). Moreover, all four transcript variants of Gs{alpha} show a high degree of sequence conservation between different species (3, 34), which would also be indicative for different functions of Gs{alpha}-L and Gs{alpha}-S. This leads to an additional explanation model for the phenotype of our patient in that not only the diminished entire activity of Gs{alpha} but also the deficiency of specific functions of Gs{alpha}-L could contribute to the phenotype. This hypothesis may be supported by the large amount of information that has been gathered during the recent decade about the unequal distribution of Gs{alpha} variants in various tissues and alterations in the relative amounts of Gs{alpha} variants, e.g. during maturation states in animals (31). Moreover, different biochemical characteristics of both variants could also be a sign for specific functions. Constitutive activity is the ability of a GPCR to adopt an active conformation in the absence of an agonist. Activating disease causing mutations have been described both in GPCRs as well as Gs{alpha} (35). But apart from these disorders, constitutive activity seems to play a role in physiological signal pathways also. Some GPCR complexes like the ß2-adrenoreceptor may have an apparent physiological constitutive activity, which is higher in nucleotide-free Gs{alpha} (36). Because Graziano et al. (37) found that Gs{alpha}-L binds GDP with a 2- to 3-fold lower affinity than Gs{alpha}-S, Gs{alpha}-L seems to be more often nucleotide free than Gs{alpha}-S and thus more frequently available to stabilize the receptor in the active state. Supporting this hypothesis, fusion proteins combined with the long variant showed in in vitro experiments a higher constitutive activity, and it has been postulated that both variants play different roles in the activation state of receptors (36, 37, 38).

In this case Gs{alpha}-L may be necessary for a constitutive activation and therefore for a basal activation of some G protein-coupled signal transduction pathways, whereas Gs{alpha}-S is rather responsible for a demand-orientated activation, which appears to interact more rapidly by agonist-bound receptor in an eukaryotic cell model (32). In our case, this may explain the development of the AHO phenotype due to only a partial loss of the long variant.


   Acknowledgments
 
We thank all family members for their participation.


   Footnotes
 
This work was supported by research grants from the German Ministry for Research and Education (Bundesministerium für Bildung und Forschung GMG 01GM0315).

Disclosure Statement: The authors have nothing to disclose.

First Published Online February 13, 2007

Abbreviations: AHO, Albright hereditary osteodystrophy; DIG, digoxigenin; fT3, free T3; fT4, free tetraiodothyronine; GPCR, G protein-coupled receptor; Gs{alpha}-L, long transcript variant; Gs{alpha}-S, short transcript variant; Nesp55, neuroendocrine secretory protein 55; PHP Ia, pseudohypoparathyroidism type Ia; SSCP, single-strand conformation polymorphism; XL{alpha}s, extra large Gs{alpha}.

Received September 28, 2006.

Accepted February 5, 2007.


   References
Top
 Abstract
 Introduction
 Patients and Methods
 Results
 Discussion
 References
 

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