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Title: Expanding GNAS-related pathology by genetic and epigenetic studies
Other Titles: Uitbreiden van GNAS-gerelateerde pathologie op basis van genetische en epigenetische studies
Authors: Izzi, Benedetta
Issue Date: 2-Dec-2011
Abstract: <!-- /* Font Definitions */@font-face {font-family:Times; panose-1:2 0 5 0 0 0 0 0 0 0; mso-font-charset:0; mso-generic-font-family:auto; mso-font-pitch:variable; mso-font-signature:3 0 0 0 1 0;}@font-face {font-family:"&#65325;&#65331; &#26126;&#26397;"; mso-font-charset:78; mso-generic-font-family:auto; mso-font-pitch:variable; mso-font-signature:-536870145 1791491579 18 0 131231 0;}@font-face {font-family:"&#65325;&#65331; &#26126;&#26397;"; mso-font-charset:78; mso-generic-font-family:auto; mso-font-pitch:variable; mso-font-signature:-536870145 1791491579 18 0 131231 0;}@font-face {font-family:Calibri; panose-1:2 15 5 2 2 2 4 3 2 4; mso-font-charset:0; mso-generic-font-family:auto; mso-font-pitch:variable; mso-font-signature:-520092929 1073786111 9 0 415 0;}@font-face {font-family:Cambria; panose-1:2 4 5 3 5 4 6 3 2 4; mso-font-charset:0; mso-generic-font-family:auto; mso-font-pitch:variable; mso-font-signature:-536870145 1073743103 0 0 415 0;}@font-face {font-family:TimesNewRomanPSMT; panose-1:0 0 0 0 0 0 0 0 0 0; mso-font-alt:"Times New Roman"; mso-font-charset:0; mso-generic-font-family:auto; mso-font-format:other; mso-font-pitch:variable; mso-font-signature:50331648 0 0 0 1 0;} /* Style Definitions */p.MsoNormal, li.MsoNormal, div.MsoNormal {mso-style-unhide:no; mso-style-qformat:yes; mso-style-parent:""; margin:0cm; margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:12.0pt; font-family:Cambria; mso-ascii-font-family:Cambria; mso-ascii-theme-font:minor-latin; mso-fareast-font-family:"&#65325;&#65331; &#26126;&#26397;"; mso-fareast-theme-font:minor-fareast; mso-hansi-font-family:Cambria; mso-hansi-theme-font:minor-latin; mso-bidi-font-family:"Times New Roman"; mso-bidi-theme-font:minor-bidi;}.MsoChpDefault {mso-style-type:export-only; mso-default-props:yes; font-family:Cambria; mso-ascii-font-family:Cambria; mso-ascii-theme-font:minor-latin; mso-fareast-font-family:"&#65325;&#65331; &#26126;&#26397;"; mso-fareast-theme-font:minor-fareast; mso-hansi-font-family:Cambria; mso-hansi-theme-font:minor-latin; mso-bidi-font-family:"Times New Roman"; mso-bidi-theme-font:minor-bidi;}@page WordSection1 {size:612.0pt 792.0pt; margin:72.0pt 90.0pt 72.0pt 90.0pt; mso-header-margin:36.0pt; mso-footer-margin:36.0pt; mso-paper-source:0;}div.WordSection1 {page:WordSection1;}--&gt;<p class="MsoNormal" style="text-align:justify;text-justify:inter-ideograph;line-height:150%"&gt;TheGNAS cluster, located on chromosome20q13 <span style="font-family:Calibri;mso-ascii-theme-font:major-latin;mso-hansi-theme-font:major-latin;mso-bidi-font-family:&amp;quot;Times New Roman&amp;quot;;color:black"&gt;gives rise to several transcripts, antisensetranscripts andnoncoding RNAs, including transcription of the gene for the stimulatory G-proteinalpha subunit (Gs<span style="font-family:Calibri;mso-ascii-theme-font:major-latin;mso-hansi-theme-font:major-latin;mso-bidi-font-family:Helvetica;color:black"&gt;&#945;<span style="font-family:Calibri;mso-ascii-theme-font:major-latin;mso-hansi-theme-font:major-latin;mso-bidi-font-family:&amp;quot;Times New Roman&amp;quot;;color:black"&gt;), which interacts with adenylyl cyclase to generate cAMP. It comprisesfour alternative first exons and promoters that splice onto exon 2 of theclassical GNAS gene, and that are alllocated in three differentially methylated regions (DMRs) namelyNESP, XL and ExonA/B. <span style="font-family:Calibri;mso-ascii-theme-font:major-latin;mso-hansi-theme-font:major-latin;mso-bidi-font-family:&amp;quot;Times New Roman&amp;quot;;color:black"&gt;The phenotypes resulting from genetic and epigenetic abnormalitiesof the GNAS region include AlbrightHereditary Osteodystrophy (AHO), pseudohypoparathyroidism types Ia (PHP-Ia) andIb (PHP-Ib), and pseudopseudohypoparathyroidism (PPHP). PHP-Ia and PPHP arecaused by inactivating GNASmutations, while PHP-Ib results from epigenetic <i style="mso-bidi-font-style:normal"&gt;GNAS mutations. All different aspects of epigenetics and with afocus on DNA methylation and the imprinted GNAScluster are reviewed in chapters 1and 3.<p class="MsoNormal" style="text-align:justify;text-justify:inter-ideograph;line-height:150%"&gt;Our research groupdeveloped a reproducible and reliable platelet-based functional Gs test, termed“platelet aggregation-inhibition test” which is based uponinhibition of platelet aggregation by cAMP after Gs stimulation. Via this test270 patients with AHO features were screened and further classified in Gs hypo-(24%), hyper- (15%) or normal function.<span style="font-family:Calibri;mso-ascii-theme-font:major-latin;mso-hansi-theme-font:major-latin;mso-bidi-font-family:Calibri;color:black"&gt; My PhD project will mainly focus on the finding of new(epi)genetic causes associated with an AHO phenotype and/or having PTHresistance in combination with a defective platelet Gs signaling pathway. In chapters 5 and 6, we have described theset up and validation of methodologies to study methylation in the different GNAS DMRs. <b style="mso-bidi-font-weight:normal"&gt;Chapter 5 reports an easy screening method to detect pronounced GNAS methylation abnormalities such as forPHP-Ib patients. Chapters 6 fully describe the optimization and validationof a quantitative methodology (Sequenom EpiTYPER mass-array) that allows thesimultaneously study of different CpGs on large samples sets. The technique wasvalidated in PHP-Ib and PHP-Ia cases to quantify their <i style="mso-bidi-font-style:normal"&gt;GNAS methylation defect. Chapter7 reports the first study of GNASCopy Number Variants (CNVs) that showedno evidence for CNVs in AHO patientswith or without abnormal Gs platelet function. PPHP patients with Gshypofunction but no GNAS codingmutation were investigated for imprinting defects in <b style="mso-bidi-font-weight:normal"&gt;chapter 8. Since AHO features are shared characteristics also withother imprinting dysorders, we have also studied other imprinting genes such asIGF2/H19, SNURF and GRB10. We found that PPHP patients have XL and IGF2hypermethylation in combination with SNURF hypomethylation. <p class="MsoNormal" style="text-align:justify;text-justify:inter-ideograph;line-height:150%"&gt;<span style="font-family:Calibri;mso-ascii-theme-font:major-latin;mso-hansi-theme-font:major-latin;mso-bidi-font-family:Calibri;color:black"&gt; <p class="MsoNormal" style="text-align:justify;text-justify:inter-ideograph;line-height:150%"&gt;<span style="font-family:Calibri;mso-ascii-theme-font:major-latin;mso-hansi-theme-font:major-latin;mso-bidi-font-family:Calibri;color:black"&gt;SinceGs hypofunction was also found in platelets from 5 spina bifida patients withAlbright’s Hereditary Osteodystrophy carrying a peculiar <i style="mso-bidi-font-style:normal"&gt;GNAS splice mutation, 96 other spina bifida cases were included forfurther (epi&#8208;)genetic GNAS studies (chapter 9). Identification of abnormalExonA/B methylation in 20% of 77 patients was the basis for further methylationstudies in SB patients. Evidence exists for a possible role for methylation inthe aetiology of NTDs (chapter 2 forreview on NTDs). We have studied for the first time DNA methylation in SB casesat genomic-scale level using the human 27K BeadChip array from Illumina thatincludes more than 27.000 CpGs. CpGs in ZNF718,KCNQ1, <i style="mso-bidi-font-style:normal"&gt;GSTT1, TP53INP1, MRI1 and <i style="mso-bidi-font-style:normal"&gt;ACOT11 were significantly hypermethylated, while the FL45964 CpGs werehypomethylated in SB patients. The genome-wide findings were replicated withthe Sequenom EpiTYPER and confirmed on a total of 101 SB patients for ACOT11, <i style="mso-bidi-font-style:normal"&gt;GSTT1 and ZNF718. The zebrafish ortholog <i style="mso-bidi-font-style:normal"&gt;ACOT11a gene was depleted in zebrafish embryos and life-screeningexperiments revealed a phenotype of neurulation defects but this deservesfurther characterization. Further studies on larger patients sets are needed tobetter characterize these preliminary findings. <span style="font-family:Calibri;mso-ascii-theme-font:major-latin;mso-hansi-theme-font:major-latin;mso-bidi-font-family:Times;color:#141413"&gt;<p class="MsoNormal" style="text-align:justify;text-justify:inter-ideograph;line-height:150%"&gt;<span style="font-family:Calibri;mso-ascii-theme-font:major-latin;mso-hansi-theme-font:major-latin;mso-bidi-font-family:Times;color:#141413"&gt; <p class="MsoNormal" style="text-align:justify;text-justify:inter-ideograph;line-height:150%"&gt;<span style="font-family:Calibri;mso-ascii-theme-font:major-latin;mso-hansi-theme-font:major-latin"&gt;In conclusion, we have expanded human GNAS-related pathology by both geneticand epigenetic studies revealing a potential relevant role of this gene in theaetiology of different pathological phenotypes, once again highlighting theextreme complexity of this fascinating, and still not completely known,imprinted gene cluster.<p class="MsoNormal" style="text-align:justify;text-justify:inter-ideograph;line-height:150%"&gt;<span style="font-family:Calibri;mso-ascii-theme-font:major-latin;mso-hansi-theme-font:major-latin;mso-bidi-font-family:Calibri;color:black"&gt;
Publication status: published
KU Leuven publication type: TH
Appears in Collections:Molecular and Vascular Biology
Pediatric Hematology & Oncology Section (-)

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