Search

Xist RNA, which is responsible for X inactivation, is a key epigenetic player in the embryogenesis of female mammals. Of the several repeats conserved in Xist RNA, the A-repeat has been shown to be essential for its silencing function in differentiating embryonic stem cells. Here, we introduced a new Xist allele into mouse that produces mutated Xist RNA lacking the A-repeat (XistCAGΔ5′). XistCAGΔ5′ RNA expressed in the embryo coated the X chromosome but failed to silence it. Although imprinted X inactivation was substantially compromised upon paternal transmission, allele-specific RNA-seq in the trophoblast revealed that XistCAGΔ5′ RNA still retained some silencing ability. Furthermore, the failure of imprinted X inactivation had more significant impacts than expected on genome-wide gene expression. It is likely that dosage compensation is required not only for equalizing X-linked gene expression between the sexes but also for proper global gene regulation in differentiated female somatic cells. [ABSTRACT FROM AUTHOR]

The dosage difference of X-linked genes between the sexes in mammals is compensated for by genetic inactivation of one of the X chromosomes in XX females. A noncoding RNA transcribed from the Xist gene at the onset of X chromosome inactivation coats the X chromosome in cis and induces chromosome-wide heterochromatinization. Here, we report a new Xist allele (XistCAG) driven by a CAG promoter, which is known to be constitutively active in many types of cells. The paternal transmission of XistCAG resulted in the preferential inactivation of the targeted paternal X (Xp) not only in the extra-embryonic but also the embryonic lineage, whereas maternal transmission ended with embryonic lethality at the early postimplantation stage with a phenotype that resembled mutant embryos carrying amaternal deficiency in Tsix, an antisense negative regulator of Xist, in both sexes. Interestingly, we found that the upregulation of XistCAG in preimplantation embryos temporally differed depending on its parental origin: its expression started at the 4- to 8-cell stages when paternally inherited, and XistCAG was upregulated at the blastocyst stage when maternally inherited. This might indicate that the Xist locus on Xp is permissive to transcription, but the Xist locus on the maternal X (Xm) is not.We extrapolated from these findings that the maternal Xist allele might manifest a chromatin structure inaccessible by transcription factors relative to the paternal allele. This might underlie the mechanism for the maternal repression of Xist at the early cleavage stage when Tsix expression has not yet occurred on Xm. [ABSTRACT FROM AUTHOR]

In germ cells, early embryos, and stem cells of animals, PIWI-interacting RNAs (piRNAs) have an important role in silencing retrotransposons, which are vicious genomic parasites, through transcriptional and post-transcriptional mechanisms. To examine whether the piRNA pathway can be used to silence genes of interest in germ cells, we have generated knock-in mice in which a foreign DNA fragment was inserted into a region generating pachytene piRNAs. The knock-in sequence was transcribed, and the resulting RNA was processed to yield piRNAs in postnatal testes. When reporter genes possessing a sequence complementary to portions of the knock-in sequence were introduced, they were greatly repressed after the time of pachytene piRNA generation. This repression mainly occurred at the post-transcriptional level, as degradation of the reporter RNAs was accelerated. Our results show that the piRNA pathway can be used as a tool for sequence-specific gene silencing in germ cells and support the idea that the piRNA generating regions serve as traps for retrotransposons, enabling the host cell to generate piRNAs against active retrotransposons. [ABSTRACT FROM AUTHOR]