A GNAS1 imprinting defect in pseudohypoparathyroidism type IB
Jie Liu, … , Leslie G. Biesecker, Lee S. Weinstein
Jie Liu, … , Leslie G. Biesecker, Lee S. Weinstein
Published November 1, 2000
Citation Information: J Clin Invest. 2000;106(9):1167-1174. https://doi.org/10.1172/JCI10431.
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Article

A GNAS1 imprinting defect in pseudohypoparathyroidism type IB

Abstract

Pseudohypoparathyroidism type IB (PHPIB) is characterized by renal resistance to parathyroid hormone (PTH) and the absence of other endocrine or physical abnormalities. Familial PHPIB has been mapped to 20q13, near GNAS1, which encodes Gsα, the G protein α-subunit required for receptor-stimulated cAMP generation. However, Gsα function is normal in blood cells from PHPIB patients, ruling out mutations within the Gsα coding region. In mice Gsα is expressed only from the maternal allele in renal proximal tubules (the site of PTH action) but is biallelically expressed in most other tissues. Studies in patients with Albright hereditary osteodystrophy suggest a similar Gsα imprinting pattern in humans. Here we identify a region upstream of the Gsα promoter that is normally methylated on the maternal allele and unmethylated on the paternal allele, but that is unmethylated on both alleles in all 13 PHPIB patients studied. Within this region is an alternative promoter and first exon (exon 1A), generating transcripts that are normally expressed only from the paternal allele, but that are biallelically expressed in PHPIB patients. Therefore, PHPIB is associated with a paternal-specific imprinting pattern of the exon 1A region on both alleles, which may lead to decreased Gsα expression in renal proximal tubules. We propose that loss of exon 1A imprinting is the cause of PHPIB.

Authors

Jie Liu, Deborah Litman, Marjorie J. Rosenberg, Shuhua Yu, Leslie G. Biesecker, Lee S. Weinstein

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RT-PCR analysis of blood RNA. (a) RT-PCR was performed on total RNA isol...
RT-PCR analysis of blood RNA. (a) RT-PCR was performed on total RNA isolated from blood of two normal subjects (N1 and N2) and two PHPIB patients (patients 1 and 3) using exon 1A-specific upstream and exon 2-specific downstream primers. Direct sequencing of RT-PCR products is shown above with the position of the polymorphic 5-bp insertion indicated by brackets. For N1 and N2, only RT-PCR products with the 5-bp insertion are identified (which for both N1 and N2 is the paternal allele; see Figure 2a). In contrast, RT-PCR products both with and without the polymorphic 5-bp insertion are amplified from patients 1 and 3, indicating biallelic expression of exon 1A mRNAs in these patients. Below each sequence are results of genotyping of subcloned products. For patient 1, clones with the 5-bp insertion are derived from the paternal allele, whereas for patient 3 these clones are derived from the maternal allele. (b) RT-PCR was performed on total RNA isolated from blood of normal subjects (N) and four PHPIB patients (patients 1, 5, 8, and 12) using NESP55-specific primers (above) or β-actin–specific primers (below). In patients 1, 5, and 8 both GNAS1 alleles are methylated in the NESP55 upstream region, whereas in patient 12 only one allele is methylated in this region (see Table 1). The presence or absence of enzyme in the RT reaction is shown above each lane.