Quantifying mRNA targeting to P bodies in living human cells reveals a dual role in mRNA decay and storage

Abstract

The 5′→3′ mRNA degradation machinery localizes to cytoplasmic P bodies (PBs), which are non-membranous structures found in all eukaryotes. While PB function has been intensively studied in yeast, less is known in mammalian cells, such as whether PB enzymes are actively engaged in mRNA degradation or if PBs serve as mRNA storage depots, particularly during cellular stress. We examined the fate of mammalian mRNAs in PBs during translational stress, and show that mRNAs accumulate within PBs during amino acid starvation. The 5′ and 3′ ends of the transcripts residing in PBs could be identified, but polyA tails were not detected. Using the MS2 mRNA-tagging system for mRNA visualization in living cells, we found that a stationary mRNA population formed in PBs during translational stress, which cleared gradually after the stress was relieved. Dcp2 knockdown experiments showed that there is constant degradation of part of the PB-associated mRNA population. This analysis demonstrates the dual role of PBs as decay sites and storage areas under regular and stress conditions.