Identificação de genes/mecanismos moleculares associados à deficiência intelectual em mulheres a partir de padrões extremos de desvio de inativação do cromossomo X

Abstract

Intellectual disability (ID) is a public health problem with high prevalence in general population (2-3%) that affects 30% more males than females. One of the factors associated with this sex ratio discrepancy is the participation of mutations in X chromosome genes. Since females present a X chromosome inactivation (XCI) mechanism to compensate the gene dosage between homo and heterogametic sexes, females carrying mutations on this chromosome may not be severely affected by ID. Thus, studies on X-linked ID (XLID) including females are rare and the knowledge about genes on the X chromosome causing ID in females is limited. Based on this, the main aim of this study was to characterize, from extreme XCI skewing, genetic alterations involved in ID in females. The XCI profile (AR and RP2 genes) was evaluated in DNA samples (peripheral blood and buccal mucosa) from 53 females with ID, and copy number variations (CNVs) in genes associated with ID were investigated through multiplex ligation-dependent probe amplification (MLPA). For the samples identified with moderate (? 80%) and extreme XCI skewing (? 90%), variants on XIST gene, the main involved in XCI, were also investigated. Besides, samples with extreme XCI skewing were submitted to comparative genomic hybridization (array-CGH) and whole exome sequencing (WES) methodologies. Four patients with moderate XCI skewing and seven patients with extreme XCI skewing were found. Among the patients with XCI skewing, ID-related changes were identified, including two pathogenic CNVs on chromosomes 3 (deletion in 3q26) and X (t(2;X)-der(2)), and four point mutations on HDAC8, NLGN4X, TAF1 and USP9X genes, two of them unpublished. In parallel, six patients with interstitial and subtelomeric CNVs were found, one of them pathogenic (22q11.21 deletion), two probably benign (RABL2B and TERT duplications) and three possibly pathogenic (SHOX duplication, SOX12 and RBFA deletions). The percentage of extreme XCI skewing observed in this study (20,7%) was significantly higher than expected in healthy females (p<0,001, ?²=18,61). The XCI assay was effective in evaluating all studied samples and allowed the identification of XCI profiles in homozygous females for the AR gene, including a patient with a DILX mutation. In addition, CNVs investigation allowed the identification of a phenotypically atypic case of 22q11.2 deletion syndrome, and of three other CNVs possibly associated with ID. In conclusion, XCI skewing proved to be an excellent tool for the characterization of (new) genes/molecular mechanisms associated with DILX in females, allowing the identification of previously unrecognized alterations responsible for ID and XCI skewing.