Project description:Using a genome-scale CRISPR knockout screening in mouse embryonic stem cells, we identify m6A RNA methylation as an important regulatory component that restricts the activity of endogenous retroviruses of the IAP (intracisternal A-particles) family. The m6A methylation of IAP mRNAs occurs on their 5’ end, is catalyzed by the complex of methyltransferase-like (METTL)3/METTL14 proteins whose depletion, along with other complex subunits, namely WTAP and ZC3H13, leads to increased IAP transcript abundance. Using auxin-dependent degron, we show that rapid removal of METTL3, METTL14 and ZC3H13 further increases IAP RNA abundance in a progressive and reversible fashion. Finally, we demonstrate that m6A RNA reduces the stability of IAP transcripts
Project description:Using a genome-scale CRISPR knockout screening in mouse embryonic stem cells, we identify m6A RNA methylation as an important regulatory component that restricts the activity of endogenous retroviruses of the IAP (intracisternal A-particles) family. The m6A methylation of IAP mRNAs occurs on their 5’ end, is catalyzed by the complex of methyltransferase-like (METTL)3/METTL14 proteins whose depletion, along with other complex subunits, namely WTAP and ZC3H13, leads to increased IAP transcript abundance. Using auxin-dependent degron, we show that rapid removal of METTL3, METTL14 and ZC3H13 further increases IAP RNA abundance in a progressive and reversible fashion. Finally, we demonstrate that m6A RNA reduces the stability of IAP transcripts
Project description:Using a genome-scale CRISPR knockout screening in mouse embryonic stem cells, we identify m6A RNA methylation as an important regulatory component that restricts the activity of endogenous retroviruses of the IAP (intracisternal A-particles) family. The m6A methylation of IAP mRNAs occurs on their 5’ end, is catalyzed by the complex of methyltransferase-like (METTL)3/METTL14 proteins whose depletion, along with other complex subunits, namely WTAP and ZC3H13, leads to increased IAP transcript abundance. Using auxin-dependent degron, we show that rapid removal of METTL3, METTL14 and ZC3H13 further increases IAP RNA abundance in a progressive and reversible fashion. Finally, we demonstrate that m6A RNA reduces the stability of IAP transcripts
Project description:Using a genome-scale CRISPR knockout screening in mouse embryonic stem cells, we identify m6A RNA methylation as an important regulatory component that restricts the activity of endogenous retroviruses of the IAP (intracisternal A-particles) family. The m6A methylation of IAP mRNAs occurs on their 5’ end, is catalyzed by the complex of methyltransferase-like (METTL)3/METTL14 proteins whose depletion, along with other complex subunits, namely WTAP and ZC3H13, leads to increased IAP transcript abundance. Using auxin-dependent degron, we show that rapid removal of METTL3, METTL14 and ZC3H13 further increases IAP RNA abundance in a progressive and reversible fashion. Finally, we demonstrate that m6A RNA reduces the stability of IAP transcripts
Project description:Zygotic genome activation (ZGA) is a critical post-fertilization step that promotes totipotency and allows different cell fates to emerge in the developing embryo. MERVL, murine endogenous retrovirus-L, is transiently upregulated at the 2-cell stage around the time of ZGA. Although MERVL expression is widely used as a marker of totipotency, the role of this retrotransposon in mouse embryogenesis remains elusive. Here, we develop methods for consistent knockdown (KD) and inactivation of interspersed copies of MERVL and show that MERVL transcripts, but not encoded retroviral proteins and their long terminal repeat (LTR) promoter that drives chimeric transcripts with host genes, is essential for accurate regulation of the host transcriptome and chromatin state during preimplantation development. Both KD and CRISPRi-based repression of MERVL result in embryonic lethality due to defects in differentiation and genomic stability. Furthermore, transcriptome and epigenome analysis revealed that loss of MERVL transcript led to retain an accessible chromatin state at, and aberrant expression of, a subset of 2-cell specific genes at mid-preimplantation stages. Taken together, our results suggest a model in which an endogenous retrovirus plays a critical role in regulating host cell fate potential.