Abstract
RNase III enzymes are fundamental to the biogenesis of microRNAs (miRNAs) and small interfering RNAs (siRNAs) in all species studied. Although alternative miRNA pathways independent of Drosha or Dicer exist, each still requires one RNase III-type enzyme. Here, we describe two strategies that marry either RNase Z or the Integrator complex with the slicing activity of Argonaute2 to generate highly functional mature miRNAs. We provide stringent validation of their RNase III independence by demonstrating efficient miRNA biogenesis and activity in Drosha and Dicer knockout cells. These data provide proof-of-principle evidence for additional mechanistic possibilities for efficient generation of small regulatory RNAs, and represent novel silencing triggers that may be exploited for technical purposes.
Publication types
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Research Support, N.I.H., Extramural
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Argonaute Proteins / deficiency
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Argonaute Proteins / genetics
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Argonaute Proteins / metabolism
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Base Sequence
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Cell Line
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DEAD-box RNA Helicases / deficiency
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DEAD-box RNA Helicases / genetics
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DEAD-box RNA Helicases / metabolism
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Endoribonucleases / deficiency
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Endoribonucleases / genetics
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Endoribonucleases / metabolism
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Gene Knockout Techniques
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HEK293 Cells
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HeLa Cells
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Herpesvirus 2, Saimiriine / genetics
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Herpesvirus 2, Saimiriine / metabolism
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Humans
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Mice
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MicroRNAs / biosynthesis*
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MicroRNAs / genetics
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RNA, Small Nuclear / genetics
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RNA, Small Nuclear / metabolism
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RNA, Viral / genetics
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RNA, Viral / metabolism
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Ribonuclease III / deficiency
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Ribonuclease III / genetics
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Ribonuclease III / metabolism*
Substances
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AGO2 protein, human
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Argonaute Proteins
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MicroRNAs
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RNA, Small Nuclear
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RNA, Viral
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Endoribonucleases
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RNase Z
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DICER1 protein, human
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DROSHA protein, human
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Ribonuclease III
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DEAD-box RNA Helicases