ABSTRACT
The Argonaute proteins (AGOs) are well known for their role in post-transcriptional gene silencing in the microRNA (miRNA) pathway. Here we show that in mouse embryonic stem cells, AGO1&2 serve additional functions that go beyond the miRNA pathway. Through the combined deletion of both Agos, we identified a specific set of genes that are uniquely regulated by AGOs but not by the other miRNA biogenesis factors. Deletion of Ago2&1 caused a global reduction of the repressive histone mark H3K27me3 due to downregulation at protein levels of Polycomb repressive complex 2 components. By integrating chromatin accessibility, prediction of transcription factor binding sites, and chromatin immunoprecipitation sequencing data, we identified the pluripotency factor KLF4 as a key modulator of AGO1&2-regulated genes. Our findings revealed a novel axis of gene regulation that is mediated by noncanonical functions of AGO proteins that affect chromatin states and gene expression using mechanisms outside the miRNA pathway.
Subject(s)
Argonaute Proteins , MicroRNAs , Animals , Argonaute Proteins/genetics , Argonaute Proteins/metabolism , Chromatin/genetics , Kruppel-Like Factor 4/genetics , Mice , MicroRNAs/genetics , MicroRNAs/metabolism , Mouse Embryonic Stem Cells/metabolism , Polycomb Repressive Complex 2/geneticsABSTRACT
Argonaute proteins (AGOs), which play an essential role in cytosolic post-transcriptional gene silencing, have been also reported to function in nuclear processes like transcriptional activation or repression, alternative splicing and, chromatin organization. As most of these studies have been conducted in human cancer cell lines, the relevance of AGOs nuclear functions in the context of mouse early embryonic development remains uninvestigated. Here, we examined a possible role of the AGO1 protein on the distribution of constitutive heterochromatin in mouse embryonic stem cells (mESCs). We observed a specific redistribution of the repressive histone mark H3K9me3 and the heterochromatin protein HP1α, away from pericentromeric regions upon Ago1 depletion. Furthermore, we demonstrated that major satellite transcripts are strongly up-regulated in Ago1_KO mESCs and that their levels are partially restored upon AGO1 rescue. We also observed a similar redistribution of H3K9me3 and HP1α in Drosha_KO mESCs, suggesting a role for microRNAs (miRNAs) in the regulation of heterochromatin distribution in mESCs. Finally, we showed that specific miRNAs with complementarity to major satellites can partially regulate the expression of these transcripts.
