Deletion of Xist repeat B disrupts cell cycle and asymmetric cell division through Usp9x hyperactivation in mice

During X-inactivation (XCI), Xist RNA establishes silencing by coating the chromosome in cis and binding diverse proteins to promote formation of a heterochromatic domain. However, Xist repeat B role beyond initiation of XCI remains unclear. Here, we find that loss of Xist repeat B in female mice allows survival and leads to a small body size persisting throughout life. Epigenetic and transcriptomic analyses reveal low levels of H3K27me3 and H2AK119ub occupancy on the X chromosome, except in certain CpG island regions, and partial reactivation of X-linked genes on the inactive X across multiple tissues. Notably, overdosage of Usp9x promotes centrosome amplification and chromosome instability, increasing the propensity of tumorigenesis. We further demonstrate that Usp9x overdosage leads to abnormalities in asymmetric cell division, thereby affecting the process of cell differentiation. Thus, Xist repeat B is necessary for gene-specific silencing during XCI maintenance and impacts cell proliferation and differentiation during development. This provides insights into repeat B importance in maintaining XCI. mRNA profiles of mouse liver, brain and primary MEF cells following Xist repeat B knockout. The mRNA-seqencing was performed in three replications.

在X染色体失活（X-inactivation, XCI）过程中，Xist RNA通过顺式包裹染色体并结合多种蛋白质，促进异染色质结构域的形成，从而建立基因沉默。然而，Xist重复序列B在X染色体失活启动之外的功能仍未明确。本研究发现，雌性小鼠中Xist重复序列B的缺失不会致死，反而会导致终生持续的小型体型。表观遗传与转录组分析结果显示，除特定CpG岛区域外，X染色体上H3K27me3与H2AK119ub的修饰水平较低，且多个组织中失活X染色体上的X连锁基因出现部分重新激活。值得注意的是，Usp9x过表达会促进中心体扩增与染色体不稳定性，提升肿瘤发生风险。本研究进一步证实，Usp9x过表达会引发不对称细胞分裂异常，进而影响细胞分化进程。综上，Xist重复序列B是X染色体失活维持过程中基因特异性沉默的必需元件，同时会影响发育过程中的细胞增殖与分化。该研究阐明了Xist重复序列B在维持X染色体失活过程中的重要作用。本数据集包含Xist重复序列B敲除后小鼠肝脏、大脑及原代小鼠胚胎成纤维细胞（primary MEF cells）的mRNA表达谱，所有样本均完成了三次重复的mRNA测序。