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Published July 7, 2016 | Accepted Version + Supplemental Material
Journal Article Open

Cutoff Suppresses RNA Polymerase II Termination to Ensure Expression of piRNA Precursors

Abstract

Small non-coding RNAs called piRNAs serve as guides for an adaptable immune system that represses transposable elements in germ cells of Metazoa. In Drosophila the RDC complex, composed of Rhino, Deadlock and Cutoff (Cuff) bind chromatin of dual-strand piRNA clusters, special genomic regions, which encode piRNA precursors. The RDC complex is required for transcription of piRNA precursors, though the mechanism by which it licenses transcription remained unknown. Here, we show that Cuff prevents premature termination of RNA polymerase II. Cuff prevents cleavage of nascent RNA at poly(A) sites by interfering with recruitment of the cleavage and polyadenylation specificity factor (CPSF) complex. Cuff also protects processed transcripts from degradation by the exonuclease Rat1. Our work reveals a conceptually different mechanism of transcriptional enhancement. In contrast to other factors that regulate termination by binding to specific signals on nascent RNA, the RDC complex inhibits termination in a chromatin-dependent and sequence-independent manner.

Additional Information

© 2016 Elsevier Inc. Received: December 28, 2015; Revised: April 4, 2016; Accepted: May 6, 2016; Published: June 9, 2016. We thank members of the A.A.A. lab for discussion. We thank Igor Antoshechkin (Caltech) for help with sequencing and Sergei Manakov for processing the RNA-seq data. E.S. was supported by a PhD fellowship of the Boehringer Ingelheim Fonds. This work was supported by grants from the NIH (R01 GM097363 and DP2 OD007371A), by the Searle Scholar and the Packard Fellowship Awards to A.A.A., by a Ellison Medical Foundation award to K.F.T., grants from the Canadian Institutes of Health Research (MOP-14409) and the Natural Sciences and Engineering Research Council of Canada (RGPIN-201) to H.D.L., a grant from the Natural Sciences and Engineering Research Council of Canada (RGPIN-435985) to C.A.S., and by funds from the Abby Rockefeller Mauze Trust and Maloris and STARR Foundations to D.J.P. Author Contributions: Experiments were designed and performed by Y.-C.A.C. piRNA-seq, total RNA-seq, and polyA RNA-seq in Figures 1 and 2A, and Figures 2C, 2D, 4B, 4C, 4E, 4F, 4H, 6A, 6B, 6E, 7A–7C, and 7E–7G, E.S. GRO-seq and chromRNA-seq in Figures 1, 2A, 2B, and 3, Y.L. Figures 2D, 4D, 4G, 5, 6C, 6D, and 7D, A.L.T. Pld promoter deletion and piRNA analysis, Figure 2A, with help from E.R. and S.V. All bioinformatic analysis was performed by M.N., and S.L. and D.J.P. modeled the Cuff structure. J.D.L., C.A.S., and H.D.L. generated the CBP80 antibody. A.A.A. and K.F.T. aided in study design and data interpretation. Y.-C.A.C. and A.A.A. wrote the paper with input from E.S., M.N., Y.L., and K.F.T.

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Accepted Version - nihms-795577.pdf

Supplemental Material - mmc1.pdf

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Created:
August 20, 2023
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October 19, 2023