Transcriptomic analysis of RTT neurons expressing miR106a-specifc sponge

The transcriptional silencing of a mammalian X chromosome is a complex but well-coordinated epigenetic process. The induction ofXistlong non-coding RNA and several protein-coding genes triggers reversible and dynamic epigenetic events to initiate, establish, and maintain X chromosome inactivation (XCI). However, small non-coding RNA function in XCI remains unidentified. Our genome-wide CRISPR/Cas9 screen in female fibroblasts examined the role of microRNA (miRNAs) in XCI. A striking finding was the identification of miR106a among the top genes from the screen. Through multiple approaches, we show that miR106a physically interacts with the conserved repeat A region in Xist. Surprisingly, this uncanonical miR106a-RepA pairing is required for the formation of N6-methyladenosine, an epigenetic RNA base modifier, which in turn stabilizes Xist.The XCI interference via miR106a inhibition has therapeutic implications for Rett syndrome (RTT) girls with a defectiveX-linked MECP2gene. Notably, the activation of silenced Mecp2 reverses the RTT-like phenotypes in adult mice1. Here, we discovered that the genetic inhibition of miR106a expressed Mecp2 from the inactive X chromosome in the brain of Tsix-Mecp2, a female symptomatic RTT murine model. We found that miR106a inhibition in Tsix-Mecp2 female mice significantly improved several facets of RTT pathology: the life span was increased, locomotor activity and exploratory behavior were improved, and apneas and breathing variabilities were diminished. Finally, miR106a-dependent MECP2 restoration in female human post-mitotic neurons normalizes morphological and functional deficits in RTT neurons. Together, our results implicate miR106a as a regulator in the transcriptional repression of the X chromosome and suggest that miR106a targeting might offer a feasible therapeutic strategy for RTT and other monogenic X-linked neurodevelopmental disorders. Human T158M neuron with Rett mutant is treated with miR inhibition, and the totally RNA was extracted from treated and control neurons. RNA libraries for RNA seq were prepared by Novogene. Sequence reads were trimmed for adaptor sequence/low quality sequence and the gene expression were then compared between the treated and control groups.