Methylation Patterns of the Novel DΜR of GNAS (GNAS-AS2) in the Pseudohypoparathyroidism 1B (iPPSD3) Subtypes

Presentation Number: SAT 359
Date of Presentation: April 1st, 2017

Patrick Hanna*1, Anne Rochtus2, Deborah Mackay3, Bruno Francou4, Jérôme Bouligant4, Anne Mantel4, Elli Anagnostou5, Elpis Vlachopapadopoulou5, Dominique Gaillard6 and Agnes Linglart7
1INSERM U1169, LE KREMLIN BICETRE, France, 2Department of Pediatrics, University Hospitals, Leuven, Belgium, 3Human Genetics and Genomic Medicine, Faculty of Medicine, University of Southampton, Southampton, United Kingdom, 4Laboratory of Molecular Genetics, Pharmacogenetics and Hormonology, University Hospital of Bicetre, LE KREMLIN BICETRE, France, 5Endocrinology Department-Department of growth and development, Children's Hospital “P. & A. Kyriakou”, ATHENS, Greece, 6service de Génétique et Biologie de la Reproduction, Hôpital Maison Blanche, Centre Hospitalier Universitaire, REMIS, France, 7, INSERM and APHP, CMR Calcium-Phosphore, LE KREMLIN BICETRE, France


Pseudohypoparathyroidism type 1B (PHP1B) (iPPSD3 according to the novel classification) is a rare disorder characterized in most patients mainly by a proximal tubule resistance to the parathyroid hormone (PTH) that manifests as hypocalcemia, hyperphosphatemia and elevated PTH. PHP1B is caused by epigenetic changes at one or several Differentially Methylated Regions (DMRs) within the GNAS locus, including loss of methylation (LOM) at the GNAS-A/B DMR. The autosomal dominant PHP1B (AD-PHP1B) patients present with a loss of imprinting (LOI) restricted to the GNAS-A/B DMR, and most of them carry a recurrent maternal deletion comprising the STX16 exons 3-9 while the sporadic patients (sporPHP1B) present with broad methylation defects. Recently, Rochtus et al identified a novel DMR within the GNAS locus (GNAS-AS2).

Objectives and patients: Characterize the methylation pattern of the GNAS-AS2-DMR in patients with AD-PHP1B and LOM at the GNAS-A/B-DMR (AD+: n=10) and in patients without (AD-: n=4) STX16 deletion, sporPHP1B (n=10) and controls (n=10). STX16 and GNAS deletion were excluded in the AD- patients by MLPA and genomic multiplex and quantitative PCR of the GNAS and the STX16 regions.


1- AD- patients showed significantly higher methylation compared to controls (respectively 38% and 22,7%; P<0.05). GNAS-AS2 was unmethylated in AD+ and sporPHP1B patients (respectively 4.0% and 2.9%).

2- Subsequently, we performed bisulfite sequencing of the GNAS-AS2-DMR. First, we identified 2 CG-rich subdomains separated by 184 bp. Second, the AD- patients displayed a unique pattern of methylation (methylated on the 1st subdomain, unmethylated on the 2nd subdomain) that was absent in controls, AD+ and sporPHP1B patients.

Conclusion: We have better refined the DMR GNAS-AS2. We have identified a subgroup of PHP1B patients who have a specific pattern of methylation/signature at the GNAS-AS2 DMR.


Nothing to Disclose: PH, AR, DM, BF, JB, AM, EA, EV, DG, AL