Abstract
Understanding the dynamics of RNA targeting to membraneless organelles is essential to disentangle their functions. Here, we investigate how P-bodies (PBs) evolve during cell-cycle progression in HEK293 cells. PB purification across the cell cycle uncovers widespread changes in their RNA content, partly uncoupled from cell-cycle-dependent changes in RNA expression. Single-molecule fluorescence in situ hybridization (FISH) shows various mRNA localization patterns in PBs peaking in G1, S, or G2, with examples illustrating the timely capture of mRNAs in PBs when their encoded protein becomes dispensable. Rather than directly reflecting absence of translation, cyclic mRNA localization in PBs can be controlled by RBPs, such as HuR in G2, and by RNA features. Indeed, while PB mRNAs are AU rich at all cell-cycle phases, they are specifically longer in G1, possibly related to post-mitotic PB reassembly. Altogether, our study supports a model where PBs are more than a default location for excess untranslated mRNAs.