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

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. • RNAs exhibit diverse spatiotemporal localization patterns at PB core and periphery • Extent of stable or transient RNA-PB interactions depends on RNA functionality • Positioning of cis -regulatory miRNA target sites influences PB interaction kinetics • PBs contribute to miRNA surveillance but less to mRNA decay Single cellular RNAs stably or transiently associate with the core or periphery of processing bodies (PBs). The kinetics, localization patterns, and enrichment of RNAs at PBs depend on RNA type and function. Although PBs may not be designated sites of mRNA degradation, they contribute to miRNA surveillance. [ABSTRACT FROM AUTHOR]