A new approach to imprinting mutation detection in GNAS by Sequenom EpiTYPER system.
BACKGROUND
Pseudohypoparathyroidism type Ib (PHPIb) results from abnormal imprinting of GNAS. Familial and sporadic forms of PHPIb have distinct GNAS imprinting patterns: familial PHPIb patients have an exon A/B-only imprinting defect and an intragenic STX16 deletion, whereas sporadic PHPIb cases have abnormal imprinting of the three differentially methylated regions (DMRs) in GNAS without the STX16 deletion. Overall GNAS methylation defects have recently been detected in some PHPIa patients.


METHODS
This study describes the first quantitative methylation analysis of multiple CpG sites for three different GNAS DMRs using the Sequenom EpiTYPER in 35 controls, 12 PHPIb patients, 2 PHPIa patients and 2 patients without parathormone (PTH) resistance but having only hypocalcemia and hyperphosphatemia.


RESULTS
All patients have GNAS methylation defects typically with NESP hypermethylation versus XL and exon A/B hypomethylation while the imprinting of SNURF/SNRPN was normal. PHPIa patients showed an abnormal methylation in the three DMRs of GNAS. For the first time, a marked abnormal GNAS methylation was also found in 2 patients without PTH resistance but having hypocalcemia and hyperphosphatemia.


CONCLUSIONS
The Sequenom EpiTYPER proves to be very sensitive in detecting DNA methylation changes. Our analysis also suggests that GNAS imprinting defects might be more frequent and diverse than previously thought.
Izzi
Benedetta
B
Center for Molecular and Vascular Biology, University of Leuven, Leuven, 3000, Belgium.
Decallonne
Brigitte
B
Devriendt
Koen
K
Bouillon
Roger
R
Vanderschueren
Dirk
D
Levtchenko
Elena
E
de Zegher
Francis
F
Van den Bruel
Annick
A
Lambrechts
Diether
D
Van Geet
Chris
C
Freson
Kathleen
K
D016022:Case-Control Studies; D002864:Chromogranins; D018899:CpG Islands / Q000235:genetics; D019175:DNA Methylation; D004252:DNA Mutational Analysis / Q000379:methods*; D005260:Female; D019205:GTP-Binding Protein alpha Subunits, Gs / Q000235:genetics*; D018392:Genomic Imprinting; D006801:Humans; D054559:Hyperphosphatemia / Q000235:genetics; D006996:Hypocalcemia / Q000235:genetics; D007962:Leukocytes / Q000378:metabolism; D008297:Male; D013058:Mass Spectrometry; D005810:Multigene Family / Q000235:genetics; D011547:Pseudohypoparathyroidism / Q000235:genetics
D002864:Chromogranins; C406732:GNAS protein, human; D019205:GTP-Binding Protein alpha Subunits, Gs