Kinetic analysis of the RNAi enzyme complex
Abstract
The siRNA-directed ribonucleoprotein complex, RISC, catalyzes target RNA cleavage in the RNA interference pathway. Here, we show that siRNA-programmed RISC is a classical Michaelis-Menten enzyme in the presence of ATP. In the absence of ATP, the rate of multiple rounds of catalysis is limited by release of the cleaved products from the enzyme. Kinetic analysis suggests that different regions of the siRNA play distinct roles in the cycle of target recognition, cleavage, and product release. Bases near the siRNA 5' end disproportionately contribute to target RNA-binding energy, whereas base pairs formed by the central and 3' regions of the siRNA provide a helical geometry required for catalysis. Finally, the position of the scissile phosphate on the target RNA seems to be determined during RISC assembly, before the siRNA encounters its RNA target.
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The Argonaute-loaded miRNA programs the gene target specificity of miRISC, as nucleotides 2-7 (the seed sequence) are mainly responsible for determining target mRNA identity through base pairing.
“The Argonaute‐loaded miRNA programs the gene target specificity of the miRISC, as nucleotides 2‐7, referred to as the miRNA “seed” sequence, are mainly responsible for determining the target mRNA identity through base pairing (Grün, Wang, Langenberger, Gunsalus, & Rajewsky, 2005; Haley & Zamore, 2004; Jackson et al., 2003; Krek et al., 2005; Lewis, Burge, & Bartel, 2005; Lewis, Shih, Jones‐Rhoades”
From microRNA strand selection: Unwinding the rules (2021) · cites this paper as Haley & Zamore, 2004
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The C. elegans heterochronic gene lin-4 encodes small RNAs with antisense complementarity to lin-14
Lee, R. C., Feinbaum, R. L., & Ambros, V. (1993) · Cell · doi:10.1016/0092-8674(93)90529-y · Cited by 12,948 · see citing works
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MicroRNAs: Target Recognition and Regulatory Functions
Bartel, D. P. (2009) · Cell · doi:10.1016/j.cell.2009.01.002 · Cited by 20,068 · see citing works
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Kertesz, M., Iovino, N., Unnerstall, U., Gaul, U., & Segal, E. (2007) · Nature Genetics · doi:10.1038/ng2135 · Cited by 2,460 · see citing works
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Schwab, R., Palatnik, J. F., Riester, M., Schommer, C., Schmid, M., & Weigel, D. (2005) · Developmental Cell · doi:10.1016/j.devcel.2005.01.018 · Cited by 1,467 · see citing works
Kawamata, T., & Tomari, Y. (2010) · Trends in Biochemical Sciences · doi:10.1016/j.tibs.2010.03.009 · Cited by 481 · see citing works
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microRNA strand selection: Unwinding the rules
Medley, J. C., Panzade, G., & Zinovyeva, A. Y. (2021) · WIREs RNA · doi:10.1002/wrna.1627 · Cited by 225 · see citing works
High-Throughput Analysis Reveals Rules for Target RNA Binding and Cleavage by AGO2
Becker, W. R., Ober-Reynolds, B., Jouravleva, K., Jolly, S. M., Zamore, P. D., & Greenleaf, W. J. (2019) · Molecular Cell · doi:10.1016/j.molcel.2019.06.012 · Cited by 161 · see citing works
Self-citationDevelopment of siRNA Payloads to Target <i>KRAS</i> -Mutant Cancer
Yuan, T. L., Fellmann, C., Lee, C.-S., Ritchie, C. D., Thapar, V., Lee, L. C., Hsu, D. J., Grace, D., Carver, J. O., Zuber, J., et al. (2014) · Cancer Discovery · doi:10.1158/2159-8290.cd-13-0900 · Cited by 113 · see citing works
- Authors
- Haley, B., & Zamore, P. D.
- Venue
- Nature Structural & Molecular Biology
- Published
- 2004-05-30
- Volume
- 11
- Issue
- 7
- Pages
- 599-606
- ISSN
- 1545-9993
- 1545-9985
- Cited by
- 454
- References
- 1
- Language
- en
- Licence
- www.springer.com/tdm (tdm)
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