Genetic Evaluation of Copy Number Variations, Loss of Heterozygosity, and Single-Nucleotide Variant Levels in Human Embryonic Stem Cells With or Without Skewed X Chromosome Inactivation

Human embryonic stem cells (hESCs) exhibiting skewed X chromosome inactivation (XCI) have been reported. The copy number variations (CNVs), loss of heterozygosity (LOH), or single-nucleotide variant (SNV) events in those epigenetically distinct cells remain unknown, and whether such genetic abnormalities will influence the XCI status of hESCs is unclear. In this study, three hESCs with skewed XCI, three with random XCI, and two male hESC lines at different passages were analyzed for CNVs and LOH levels using a high-resolution genotyping microarray. Whole-exome sequencing was used to investigate the potentially damaging SNVs. On average, 17.6 CNVs and 5.3 cases of LOH were identified in the skewed hESCs, which were similar to the rates observed in random hESCs. Five recurrent CNV regions were uniquely identified in the skewed hESCs, but all of them were considered polymorphisms. With the exception of a nongenic CNV, no additional CNVs were detected on the X chromosome in the skewed hESCs. Although the XCI status in two hESC lines was observed to be changed from random to skewed, no significant CNV difference was identified before and after the XCI change. SNV analysis indicated that normal alleles are maintained for most genes within copy-neutral LOH regions. Three types of expression patterns were observed in heterozygous alleles, and the damaging SNVs in skewed hESCs favored the expression of the wild-type alleles. In conclusion, in the present study, we did not find genetic differences in the CNV and LOH levels between hESCs with and without skewed XCI. Wild-type allele expression in the presence of damaging SNVs on the X chromosome in skewed hESCs might alleviate adverse effects in those hESCs.