The N Domain of Argonaute Drives Duplex Unwinding During RISC Assembly

Overview

Journal Nat Struct Mol Biol

Specialties Cell Biology
Molecular Biology

Date 2012 Jan 12

PMID 22233755

Citations 148

Authors

Pieter Bas Kwak

Yukihide Tomari

Affiliations

Soon will be listed here.

Abstract

Small RNAs, such as microRNAs and small interfering RNAs, act through Argonaute (Ago) proteins as a part of RNA-induced silencing complexes (RISCs). To make RISCs, Ago proteins bind and subsequently unwind small RNA duplexes, finally leaving one strand stably incorporated. Here we identified the N domain of human AGO2 as the initiator of duplex unwinding during RISC assembly. We discovered that a functional N domain is strictly required for small RNA duplex unwinding but not for precedent duplex loading or subsequent target cleavage. We postulate that RISC assembly is tripartite, comprising (i) RISC loading, whereby Ago undergoes conformational opening and loads a small RNA duplex, forming pre-RISC; (ii) wedging, whereby the end of the duplex is pried open through active wedging by the N domain, in preparation for unwinding; and (iii) unwinding, whereby the passenger strand is removed through slicer-dependent or slicer-independent unwinding, forming mature RISC.

Citing Articles

Structural and mechanistic insights into the activation of a short prokaryotic argonaute system from archaeon Sulfolobus islandicus.

Dai Z, Chen Y, Guan Z, Chen X, Tan K, Yang K Nucleic Acids Res. 2025; 53(3).

PMID: 39898546 PMC: 11788926. DOI: 10.1093/nar/gkaf059.

A guide to the biogenesis and functions of endogenous small non-coding RNAs in animals.

Jouravleva K, Zamore P Nat Rev Mol Cell Biol. 2025; .

PMID: 39856370 DOI: 10.1038/s41580-024-00818-9.

piRNA Defense Against Endogenous Retroviruses.

Abajorga M, Yurkovetskiy L, Luban J Viruses. 2024; 16(11).

PMID: 39599869 PMC: 11599104. DOI: 10.3390/v16111756.

The role of prokaryotic argonautes in resistance to type II topoisomerases poison ciprofloxacin.

Galivondzhyan A, Sutormin D, Panteleev V, Kulbachinskiy A, Severinov K Biochem Soc Trans. 2024; 52(5):2157-2166.

PMID: 39446311 PMC: 11555693. DOI: 10.1042/BST20240094.

Efflux ABC transporters in drug disposition and their posttranscriptional gene regulation by microRNAs.

Wang Y, Tu M, Yu A Front Pharmacol. 2024; 15:1423416.

PMID: 39114355 PMC: 11303158. DOI: 10.3389/fphar.2024.1423416.

References

Carthew R, Sontheimer E . Origins and Mechanisms of miRNAs and siRNAs. Cell. 2009; 136(4):642-55. PMC: 2675692. DOI: 10.1016/j.cell.2009.01.035. View

Kawamata T, Seitz H, Tomari Y . Structural determinants of miRNAs for RISC loading and slicer-independent unwinding. Nat Struct Mol Biol. 2009; 16(9):953-60. DOI: 10.1038/nsmb.1630. View

Rand T, Petersen S, Du F, Wang X . Argonaute2 cleaves the anti-guide strand of siRNA during RISC activation. Cell. 2005; 123(4):621-9. DOI: 10.1016/j.cell.2005.10.020. View

Kawamata T, Tomari Y . Making RISC. Trends Biochem Sci. 2010; 35(7):368-76. DOI: 10.1016/j.tibs.2010.03.009. View

Boland A, Huntzinger E, Schmidt S, Izaurralde E, Weichenrieder O . Crystal structure of the MID-PIWI lobe of a eukaryotic Argonaute protein. Proc Natl Acad Sci U S A. 2011; 108(26):10466-71. PMC: 3127882. DOI: 10.1073/pnas.1103946108. View

Nykanen A, Haley B, Zamore P . ATP requirements and small interfering RNA structure in the RNA interference pathway. Cell. 2001; 107(3):309-21. DOI: 10.1016/s0092-8674(01)00547-5. View

Wang Y, Juranek S, Li H, Sheng G, Wardle G, Tuschl T . Nucleation, propagation and cleavage of target RNAs in Ago silencing complexes. Nature. 2009; 461(7265):754-61. PMC: 2880917. DOI: 10.1038/nature08434. View

Ma J, Ye K, Patel D . Structural basis for overhang-specific small interfering RNA recognition by the PAZ domain. Nature. 2004; 429(6989):318-322. PMC: 4700412. DOI: 10.1038/nature02519. View

Waterhouse A, Procter J, Martin D, Clamp M, Barton G . Jalview Version 2--a multiple sequence alignment editor and analysis workbench. Bioinformatics. 2009; 25(9):1189-91. PMC: 2672624. DOI: 10.1093/bioinformatics/btp033. View

10.

Khvorova A, Reynolds A, Jayasena S . Functional siRNAs and miRNAs exhibit strand bias. Cell. 2003; 115(2):209-16. DOI: 10.1016/s0092-8674(03)00801-8. View

11.

Frank F, Sonenberg N, Nagar B . Structural basis for 5'-nucleotide base-specific recognition of guide RNA by human AGO2. Nature. 2010; 465(7299):818-22. DOI: 10.1038/nature09039. View

12.

Hutvagner G, Simard M . Argonaute proteins: key players in RNA silencing. Nat Rev Mol Cell Biol. 2007; 9(1):22-32. DOI: 10.1038/nrm2321. View

13.

Liu Y, Ye X, Jiang F, Liang C, Chen D, Peng J . C3PO, an endoribonuclease that promotes RNAi by facilitating RISC activation. Science. 2009; 325(5941):750-3. PMC: 2855623. DOI: 10.1126/science.1176325. View

14.

Meister G, Landthaler M, Patkaniowska A, Dorsett Y, Teng G, Tuschl T . Human Argonaute2 mediates RNA cleavage targeted by miRNAs and siRNAs. Mol Cell. 2004; 15(2):185-97. DOI: 10.1016/j.molcel.2004.07.007. View

15.

LARKIN M, Blackshields G, Brown N, Chenna R, McGettigan P, McWilliam H . Clustal W and Clustal X version 2.0. Bioinformatics. 2007; 23(21):2947-8. DOI: 10.1093/bioinformatics/btm404. View

16.

Parker J, Roe S, Barford D . Structural insights into mRNA recognition from a PIWI domain-siRNA guide complex. Nature. 2005; 434(7033):663-6. PMC: 2938470. DOI: 10.1038/nature03462. View

17.

Kim V, Han J, Siomi M . Biogenesis of small RNAs in animals. Nat Rev Mol Cell Biol. 2009; 10(2):126-39. DOI: 10.1038/nrm2632. View

18.

Iki T, Yoshikawa M, Nishikiori M, Jaudal M, Matsumoto-Yokoyama E, Mitsuhara I . In vitro assembly of plant RNA-induced silencing complexes facilitated by molecular chaperone HSP90. Mol Cell. 2010; 39(2):282-91. DOI: 10.1016/j.molcel.2010.05.014. View

19.

Johnston M, Geoffroy M, Sobala A, Hay R, Hutvagner G . HSP90 protein stabilizes unloaded argonaute complexes and microscopic P-bodies in human cells. Mol Biol Cell. 2010; 21(9):1462-9. PMC: 2861606. DOI: 10.1091/mbc.e09-10-0885. View

20.

Song J, Smith S, Hannon G, Joshua-Tor L . Crystal structure of Argonaute and its implications for RISC slicer activity. Science. 2004; 305(5689):1434-7. DOI: 10.1126/science.1102514. View