To Degrade a MicroRNA, Destroy Its Argonaute Protein

Han et al., 2020Han J. LaVigne C.A. Jones B.T. Zhang H. Gillett F. Mendell J.T. A ubiquitin ligase mediates target-directed microRNA decay independently of tailing and trimming.Science. 2020; 370: eabc9546https://doi.org/10.1126/science.abc9546Crossref PubMed Scopus (31) Google Scholar Shi 2020Shi C.Y. Kingston E.R. Kleaveland B. Lin D.H. Stubna M.W. Bartel D.P. The ZSWIM8 degradation.Science. eabc9359https://doi.org/10.1126/science.abc9359Crossref report that the E3 senses when an RNA Argonaute protein-bound are extensively base paired directs destruction by proteasome. result is degradation microRNA. Animal microRNAs (miRNAs) rarely pair with their targets outside miRNA's seven-nucleotide seed sequence (miRNA nucleotides g2–8), a binding strategy balances target finding, affinity, specificity. Ensconced in proteins, miRNAs typically long lived. first hint complementary RNAs can trigger miRNA turnover was observation fully 2?-O-methyl oligonucleotides irreversibly inactivate Argonaute-bound small mammalian cultured cells vivo nematodes. Subsequently, artificial mimics viral non-coding extensive complementarity to endogenous were found miRNA, process now called (TDMD) (Ameres 2010Ameres S.L. Horwich M.D. Hung J.H. Xu Ghildiyal M. Weng Z. Zamore P.D. Target RNA-directed trimming silencing RNAs.Science. 2010; 328: 1534-1539Crossref (377) Scholar; Cazalla 2010Cazalla D. Yario T. Steitz J.A. Down-regulation host Herpesvirus saimiri noncoding RNA.Science. 1563-1566Crossref (352) Scholar). Now, TDMD requires ZSWIM8, associated proteolysis, not turnover. elicits its guide binds highly RNA. Strong substrates have 5? 3? often intervening unpaired region best-studied triggers Cyrano, which destabilizes miR-7, Nrep, miR-29. searched for proteins pathway using genome-wide, Cas9-sgRNA screens cell lines naturally expressing Cyrano. Each study employed miR-7-repressed reporter mRNA encoding fluorescent protein. Loss increased miR-7 abundance, thereby decreasing fluorescence. This led both groups find ZSWIM8. When protein housing instructs proteosome degrade Argonaute, miRNA. Thus, appears destroy indirectly, depriving them Argonaute. Notably, ZSWIM8-mediated has deep evolutionary roots: loss orthologs flies worms stabilizes specific miRNAs, revealing dozens new examples being used decrease half-lives. How does signal promote degradation? Both suggest change Argonaute’s conformation, allowing bind. brings it E2 ubiquitin-conjugating enzyme then polyubiquitinylates conserved lysine (Figure 1). Addition multiple ubiquitins marks Presumably, protective partner leads rapid naked Structures human AGO2 reveal miRNA-target pairing dislodges end from PAZ domain (Sheu-Gruttadauria 2019Sheu-Gruttadauria Pawlica P. Klum S.M. Wang S. T.A. Schirle Oakdale N.T. MacRae I.J. Structural Basis Target-Directed MicroRNA Degradation.Mol. Cell. 2019; 75: 1243-1255.e7Abstract Full Text PDF (60) Scholar), idea proposed 15 years ago explain why adds surprisingly little additional energy. Concomitantly, central cleft becomes wider as between increases. These two conformational changes—an empty widened cleft—provide landmarks might recognize signaling “Tough decoys” designed be transcribed polymerase III fold into pre-miRNA-like structure enables nuclear export; stem loop contains single-stranded, miRNA-binding sequences. In mouse studies tough decoys exquisitely sensitive position identity target-miRNA mismatches. discriminate among, example, decoy bound nine let-7 paralogs, efficiently degrading loaded let-7a, let-7f, miR-98, let-7c, let-7d, let-7e, but let-7g, let-7b, or let-7i? vitro vivo, accompanied addition non-templated uridines adenosines (tailing) 3?-to-5? exonucleolytic shortening (trimming). co-occurrence TDMD, together many uridylation controlling stability, suggested TDMD. show tailing-and-trimming separable: tailed trimmed ZSWIM8-deficient cells; synthetic protected terminal group, nonetheless destroyed via ZSWIM8-dependent presence demote subsidiary role, proteolysis explaining most, if all, Several biological observations argue additional, ZSWIM8-independent pathways regulate stability animals. arthropods, siRNAs 2?-O-methylated, Ago2, RNAs. Ago2-bound unstable mutant Hen1, 3?-terminal, whereas insensitive 3?-terminal modification status (Han siRNA 2?-O-methylation consequence: reactivation some somatic transposons normally silenced siRNAs. C. elegans henn1 mutants, several classes methylated become unstable, including 22G- 26G-RNAs, unique nematodes (Kamminga 2012Kamminga L.M. van Wolfswinkel J.C. Luteijn M.J. Kaaij L.J. Bagijn M.P. Sapetschnig A. Miska E.A. Berezikov E. Ketting R.F. Differential impact HEN1 homolog HENN-1 on 21U 26G germline Caenorhabditis elegans.PLoS Genet. 2012; 8: e1002702Crossref (69) piRNAs, only animals, also 2?-O-methylated Hen1. piRNAs live sea triggers: piRNA production transposon require bind hen1 re-express (Wang 2016Wang Ma Niu K. Xiao Y. Wu X. Pan Zhao Liu N. Antagonistic roles Nibbler Hen1 modulating 3¢ ends Drosophila.Development. 2016; 143: 530-539Crossref (39) (Montgomery 2012Montgomery Rim Y.S. Dowen R.H. Phillips C.M. Fischer S.E. Ruvkun G. PIWI specifically ortholog henn-1.PLoS e1002616Crossref (87) fish 2010Kamminga den Broeder Redl Roovers E.F. Ladurner required oocyte development zebrafish.EMBO 29: 3688-3700Crossref (114) mice (Lim 2015Lim Qu Z.P. Kortschak R.D. Lawrence D.M. Geoghegan Hempfling A.L. Bergmann Goodnow C.C. Ormandy C.J. Wong L. al.HENMT1 Stability Are Required Adult Male Germ Cell Transposon Repression Define Spermatogenic Program Mouse.PLoS 2015; 11: e1005620Crossref (62) Moreover, recent Henmt1 mutants subset unmethylated suggesting rather than they (“PIWI” proteins), targeted destruction. disparate mechanisms exploit distinct differences entire length Identifying what “sees” encounters likely inform our understanding pathways. P.D.Z. member advisory board Molecular inventor patents filed University Massachusetts describing use inhibit function vivo.