Mechanisms of X-linked disorders’ expression in females (modeled with Hemophilia carriers). X-Inactivation Skewing: causes, biphasic response and phenotype vs tissue specificity.

As an X-linked recessive disease, hemophilia manifests in the hemizygous male and not in the heterozygous female. However, there are several reports of female carriers with hemophilia as a consequence of extreme skewed inactivation of the X chromosome over the normal F8 or F9 allele. More rarely, there are women with hemophilia with a homozygous (by consanguinity) or double heterozygous genotypes (two different variants), and phenocopies, women with a phenotype similar to hemophilia but due to other coagulopathies. Our work focuses on the large population of HA/HB carriers diagnosed by F8/F9 genotyping with clinical and biochemical phenotypic data. This population is an ideal model to investigate the expression of recessive X-linked diseases in women. In this population of carriers, cases of women with clinical expression of hemophilia (symptomatic) stand out. We and others observed that most heterozygous carriers with a severe phenotype show extremely skewed X-inactivation in peripheral blood leukocytes (X-inactivation pattern, XIP>90%). These studies were performed using a XIP measurement system, on Xq and Xp, designed by us in collaboration, combining the AR and RP2 systems. Drawing on the valuable population of women with extreme biased inactivation from all backgrounds (including symptomatic carriers of hemophilia and Duchenne muscular dystrophy), we focused on investigating the causes of skewing and its dynamics in embryonic development and in adult life in different organs/tissues. These case/control studies (extremely-skewed vs balanced XIP) are performed against the exposure of the X-chromosome genotype. Our hypothesis is a genetic failure either in the inactivation process per se, or affecting a cellular function that block or slow the proliferation of one of the X-inactivation mosaic clones. This research has started with conventional studies of SNPs variants in the X-inactivation center (XIC). For the necessary massive screening of variants in the X-chromosome that this investigation of the causes of XIP skewing implies, we are applying SNP-CGH microarrays and new generation sequencing techniques such as exomes, etc.