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Updated: Dec 20, 2025

Biotin-based Pulldown Assay to Validate mRNA Targets of Cellular miRNAs
Published on: June 12, 2018
Base-pair conformational switch modulates miR-34a targeting of Sirt1 mRNA
Lorenzo Baronti1, Ileana Guzzetti1, Parisa Ebrahimi2
1Department of Medical Biochemistry and Biophysics, Karolinska Institute, Stockholm, Sweden.
Abstract:
MicroRNAs (miRNAs) regulate the levels of translation of messenger RNAs (mRNAs). At present, the major parameter that can explain the selection of the target mRNA and the efficiency of translation repression is the base pairing between the 'seed' region of the miRNA and its counterpart mRNA1. Here we use R1ρ relaxation-dispersion nuclear magnetic resonance2 and molecular simulations3 to reveal a dynamic switch-based on the rearrangement of a single base pair in the miRNA-mRNA duplex-that elongates a weak five-base-pair seed to a complete seven-base-pair seed. This switch also causes coaxial stacking of the seed and supplementary helix fitting into human Argonaute 2 protein (Ago2), reminiscent of an active state in prokaryotic Ago4,5. Stabilizing this transient state leads to enhanced repression of the target mRNA in cells, revealing the importance of this miRNA-mRNA structure. Our observations tie together previous findings regarding the stepwise miRNA targeting process from an initial 'screening' state to an 'active' state, and unveil the role of the RNA duplex beyond the seed in Ago2.
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