Structure-function analysis of microRNA 3′-end trimming by Nibbler Journal Article
| Authors: | Xie, W.; Sowemimo, I.; Hayashi, R.; Wang, J.; Burkard, T. R.; Brennecke, J.; Ameres, S. L.; Patel, D. J. |
| Article Title: | Structure-function analysis of microRNA 3′-end trimming by Nibbler |
| Abstract: | Nibbler (Nbr) is a 3′-to-5′ exoribonuclease whose catalytic 3′-end trimming activity impacts microRNA (miRNA) and PIWI-interacting RNA (piRNA) biogenesis. Here, we report on structural and functional studies to decipher the contributions of Nbr’s N-terminal domain (NTD) and exonucleolytic domain (EXO) in miRNA 3′-end trimming. We have solved the crystal structures of the NTD core and EXO domains of Nbr, both in the apo-state. The NTD-core domain of Aedes aegypti Nbr adopts a HEAT-like repeat scaffold with basic patches constituting an RNA-binding surface exhibiting a preference for binding double-strand RNA (dsRNA) over single-strand RNA (ssRNA). Structure-guided functional assays in Drosophila S2 cells confirmed a principal role of the NTD in exonucleolytic miRNA trimming, which depends on basic surface patches. Gain-of-function experiments revealed a potential role of the NTD in recruiting Nbr to Argonaute-bound small RNA substrates. The EXO domain of A. aegypti and Drosophila melanogaster Nbr adopt a mixed α/β-scaffold with a deep pocket lined by a DEDDy catalytic cleavage motif. We demonstrate that Nbr’s EXO domain exhibits Mn2+-dependent ssRNA-specific 3′-to-5′ exoribonuclease activity. Modeling of a 3′ terminal Uridine into the catalytic pocket of Nbr EXO indicates that 2′-O-methylation of the 3′-U would result in a steric clash with a tryptophan side chain, suggesting that 2′-O-methylation protects small RNAs from Nbr-mediated trimming. Overall, our data establish that Nbr requires its NTD as a substrate recruitment platform to execute exonucleolytic miRNA maturation, catalyzed by the ribonuclease EXO domain. © 2020 National Academy of Sciences. All rights reserved. |
| Keywords: | exoribonuclease; heat repeats; microrna trimming; nibbler |
| Journal Title: | Proceedings of the National Academy of Sciences of the United States of America |
| Volume: | 117 |
| Issue: | 48 |
| ISSN: | 0027-8424 |
| Publisher: | National Academy of Sciences |
| Date Published: | 2020-12-01 |
| Start Page: | 30370 |
| End Page: | 30379 |
| Language: | English |
| DOI: | 10.1073/pnas.2018156117 |
| PUBMED: | 33199607 |
| PROVIDER: | scopus |
| PMCID: | PMC7720153 |
| DOI/URL: | |
| Notes: | Article -- Export Date: 4 January 2021 -- Source: Scopus |
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