title: Argonaute-bound small RNAs from promoter-proximal RNA polymerase II.
abstract: Argonaute (Ago) proteins mediate posttranscriptional gene repression by binding guide miRNAs to regulate targeted RNAs. To confidently assess Ago-bound small RNAs, we adapted a mouse embryonic stem cell system to express a single epitope-tagged Ago protein family member in an inducible manner. Here, we report the small RNA profile of Ago-deficient cells and show that Ago-dependent stability is a common feature of mammalian miRNAs. Using this criteria and immunopurification, we identified an Ago-dependent class of noncanonical miRNAs derived from protein-coding gene promoters, which we name transcriptional start site miRNAs (TSS-miRNAs). A subset of promoter-proximal RNA polymerase II (RNAPII) complexes produces hairpin RNAs that are processed in a DiGeorge syndrome critical region gene 8 (Dgcr8)/Drosha-independent but Dicer-dependent manner. TSS-miRNA activity is detectable from endogenous levels and following overexpression of mRNA constructs. Finally, we present evidence of differential expression and conservation in humans, suggesting important roles in gene regulation.
authors.affiliation: David H. Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
authors.forename: Jesse R
authors.initials: JR
authors.lastname: Zamudio
authors.affiliation: David H. Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
authors.forename: Timothy J
authors.initials: TJ
authors.lastname: Kelly
authors.affiliation: David H. Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA. Electronic address: sharppa@mit.edu.
authors.forename: Phillip A
authors.initials: PA
authors.lastname: Sharp
mesh_terms: D000818:Animals; D060565:Argonaute Proteins; D001483:Base Sequence; D039223:Cleavage And Polyadenylation Specificity Factor / Q000378:metabolism; D053595:Embryonic Stem Cells / Q000378:metabolism; D005786:Gene Expression Regulation; D005821:Genetic Techniques; D006801:Humans; D051379:Mice; D011401:Promoter Regions, Genetic; D012319:RNA Polymerase II / Q000378:metabolism*; D058727:RNA, Small Untranslated / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D061805:Transcription Elongation, Genetic; D024363:Transcription Initiation Site
chemical_list: C526735:Ago2 protein, mouse; D060565:Argonaute Proteins; D039223:Cleavage And Polyadenylation Specificity Factor; D058727:RNA, Small Untranslated; D012319:RNA Polymerase II