Dynamic Recruitment of Single RNAs to Processing Bodies Depends on RNA Functionality

Sethuramasundaram Pitchiaya; Marcio D A Mourao; Ameya P Jalihal; Lanbo Xiao; Xia Jiang; Arul M Chinnaiyan; Santiago Schnell; Nils G Walter

doi:10.1016/j.molcel.2019.03.001

Dynamic Recruitment of Single RNAs to Processing Bodies Depends on RNA Functionality

Mol Cell. 2019 May 2;74(3):521-533.e6. doi: 10.1016/j.molcel.2019.03.001. Epub 2019 Apr 2.

Authors

Sethuramasundaram Pitchiaya¹, Marcio D A Mourao², Ameya P Jalihal³, Lanbo Xiao⁴, Xia Jiang⁴, Arul M Chinnaiyan⁵, Santiago Schnell⁶, Nils G Walter⁷

Affiliations

¹ Single Molecule Analysis Group, Department of Chemistry, University of Michigan, Ann Arbor, MI 48109-1055, USA; Michigan Center for Translational Pathology, University of Michigan Medical School, Ann Arbor, MI 48109-1055, USA; Department of Pathology, University of Michigan, Ann Arbor, MI 48109, USA; Comprehensive Cancer Center, University of Michigan, Ann Arbor, MI 48109, USA. Electronic address: sethu@umich.edu.
² Department of Molecular and Integrative Physiology, University of Michigan Medical School, Ann Arbor, MI 48109-1055, USA; Consulting for Statistics, Computing and Analytics Research, University of Michigan, Ann Arbor, MI 48109-1055, USA.
³ Single Molecule Analysis Group, Department of Chemistry, University of Michigan, Ann Arbor, MI 48109-1055, USA.
⁴ Michigan Center for Translational Pathology, University of Michigan Medical School, Ann Arbor, MI 48109-1055, USA.
⁵ Michigan Center for Translational Pathology, University of Michigan Medical School, Ann Arbor, MI 48109-1055, USA; Department of Pathology, University of Michigan, Ann Arbor, MI 48109, USA; Comprehensive Cancer Center, University of Michigan, Ann Arbor, MI 48109, USA; Howard Hughes Medical Institute, University of Michigan, Ann Arbor, MI 48109, USA.
⁶ Department of Molecular and Integrative Physiology, University of Michigan Medical School, Ann Arbor, MI 48109-1055, USA.
⁷ Single Molecule Analysis Group, Department of Chemistry, University of Michigan, Ann Arbor, MI 48109-1055, USA. Electronic address: nwalter@umich.edu.

Abstract

Cellular RNAs often colocalize with cytoplasmic, membrane-less ribonucleoprotein (RNP) granules enriched for RNA-processing enzymes, termed processing bodies (PBs). Here we track the dynamic localization of individual miRNAs, mRNAs, and long non-coding RNAs (lncRNAs) to PBs using intracellular single-molecule fluorescence microscopy. We find that unused miRNAs stably bind to PBs, whereas functional miRNAs, repressed mRNAs, and lncRNAs both transiently and stably localize within either the core or periphery of PBs, albeit to different extents. Consequently, translation potential and 3' versus 5' placement of miRNA target sites significantly affect the PB localization dynamics of mRNAs. Using computational modeling and supporting experimental approaches, we show that partitioning in the PB phase attenuates mRNA silencing, suggesting that physiological mRNA turnover occurs predominantly outside of PBs. Instead, our data support a PB role in sequestering unused miRNAs for surveillance and provide a framework for investigating the dynamic assembly of RNP granules by phase separation at single-molecule resolution.

Keywords: RNP granules; long non-coding RNAs; mRNAs; microRNAs; phase separation; processing bodies; single-molecule microscopy.

Publication types

Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

Cytoplasmic Granules / genetics
Gene Silencing
HeLa Cells
Humans
MicroRNAs / genetics*
RNA Processing, Post-Transcriptional / genetics
RNA, Long Noncoding / genetics*
RNA, Messenger / genetics*
RNA, Untranslated / genetics
Ribonucleoproteins / genetics*
Single Molecule Imaging

Substances

MicroRNAs
RNA, Long Noncoding
RNA, Messenger
RNA, Untranslated
Ribonucleoproteins

Abstract

Publication types

MeSH terms

Substances

Grants and funding