mRNA structural dynamics shape Argonaute-target interactions

TY - JOUR

T1 - mRNA structural dynamics shape Argonaute-target interactions

AU - Ruijtenberg, Suzan

AU - Sonneveld, Stijn

AU - Cui, Tao Ju

AU - Logister, Ive

AU - de Steenwinkel, Dion

AU - Xiao, Yao

AU - MacRae, Ian J

AU - Joo, Chirlmin

AU - Tanenbaum, Marvin E

PY - 2020/9

Y1 - 2020/9

N2 - Small interfering RNAs (siRNAs) promote RNA degradation in a variety of processes and have important clinical applications. siRNAs direct cleavage of target RNAs by guiding Argonaute2 (AGO2) to its target site. Target site accessibility is critical for AGO2-target interactions, but how target site accessibility is controlled in vivo is poorly understood. Here, we use live-cell single-molecule imaging in human cells to determine rate constants of the AGO2 cleavage cycle in vivo. We find that the rate-limiting step in mRNA cleavage frequently involves unmasking of target sites by translating ribosomes. Target site masking is caused by heterogeneous intramolecular RNA-RNA interactions, which can conceal target sites for many minutes in the absence of translation. Our results uncover how dynamic changes in mRNA structure shape AGO2-target recognition, provide estimates of mRNA folding and unfolding rates in vivo, and provide experimental evidence for the role of mRNA structural dynamics in control of mRNA-protein interactions.

AB - Small interfering RNAs (siRNAs) promote RNA degradation in a variety of processes and have important clinical applications. siRNAs direct cleavage of target RNAs by guiding Argonaute2 (AGO2) to its target site. Target site accessibility is critical for AGO2-target interactions, but how target site accessibility is controlled in vivo is poorly understood. Here, we use live-cell single-molecule imaging in human cells to determine rate constants of the AGO2 cleavage cycle in vivo. We find that the rate-limiting step in mRNA cleavage frequently involves unmasking of target sites by translating ribosomes. Target site masking is caused by heterogeneous intramolecular RNA-RNA interactions, which can conceal target sites for many minutes in the absence of translation. Our results uncover how dynamic changes in mRNA structure shape AGO2-target recognition, provide estimates of mRNA folding and unfolding rates in vivo, and provide experimental evidence for the role of mRNA structural dynamics in control of mRNA-protein interactions.

KW - Argonaute Proteins/metabolism

KW - Cell Line

KW - HEK293 Cells

KW - Humans

KW - Nucleic Acid Conformation

KW - RNA Cleavage

KW - RNA Folding

KW - RNA, Messenger/chemistry

KW - Ribosomes/metabolism

U2 - 10.1038/s41594-020-0461-1

DO - 10.1038/s41594-020-0461-1

M3 - Article

C2 - 32661421

SN - 1545-9993

VL - 27

SP - 790

EP - 801

JO - Nature Structural & Molecular Biology

JF - Nature Structural & Molecular Biology

IS - 9

ER -