EXPLORING THE CAUSES OF SKEWED X-CHROMOSOME INACTIVATION: XIST GENETIC ANALYSIS OF CASES AND CONTROLS.

RADIC CP; MORA EMILCE; MARCHIONE VD; ABELLEYRO MM; GILIBERTO F; SZIJAN I; ROSSETTI L; NEME D; DE BRASI CARLOS

Orlando

Congreso; The XXXII International Congress of the World Federation of Hemophilia; 2016

Random X-chromosome inactivation (XCI) of one of the two X-chromosomes in females achieves dosage equivalency for X-linked genes with males XY. Skewed XCI is a strong deviation of 50% and among others causes may be associated with deletions in XIST (X-inactivation specific transcript), non coding gene responsible for initiation, spreading and maintenance of XCI. The presence of genetic variants as SNP (Single Nucleotide Polymorphism) may modify XIST expression and/or function impacting its allelic ability to inactivate. This work is aimed to study the association between those variants with skewed XCI. Materials and Methods: The studied population included 22 women, 11 cases with skewed XCI (90-100%) and 11 controls with fairly random XCI (50-55%). Cases: carriers (symptomatic and asymptomatic) and non-carriers of Haemophilia A and Duchenne Muscular Dystrophy. A comprehensive genetic variant screening of XIST was performed by CSGE (conformation sensitive gel electrophoresis) and Sanger sequencing. Most relevant sequences of XIST were studied: the Promoter (8 amplicons) and the exons 2 to 5 with its splicing consensus were entirely PCR-amplified and screened (4 amplicons: 64, 137, 209 and 164pb) and exon 1 (11372pb) and 6 (7325pb) included only regions with SNPs with known allelic frequencies (8 amplicons).Results: The analysis of 440 amplicons showed no new mutations but revealed 7 allelic variants from the studied 80 XIST SNPs. Statistical analysis: OR (odds ratio) associated with 2 exon 6 SNPs´ (rs1794213 and rs1620574) suggested a protective effect (0.30≤OR≤0.33); a SNP in the Promoter (rs41305409) was neutral (OR=1) and 4 SNPs, 3 in exon 6 (rs16992443, rs16992436 and rs16992442) and 1 in exon 1 (rs6527), associated with skewed XCI (2.6≤OR≤3.8). Perhaps because of samples´ sizes, none of the differences reached statistical significance (P44) and extending the targeted regions to the entire exons 1 and 6, and deep intronic sequences.