RPS28B mRNA acts as a scaffold promoting cis-translational interaction of proteins driving P-body assembly

Abstract

P-bodies (PBs) are cytoplasmic mRNA-protein (mRNP) granules conserved throughout eukaryotes which are implicated in the repression, storage and degradation of mRNAs. PB assembly is driven in part by proteins with self-interacting and low-complexity protein domains.Non-translating mRNA is also required for PB assembly, however no studies to date have explored whether particular mRNA transcripts are more critical than others in facilitating PB assembly. A previous genome-wide microscopy screen in yeast revealed that rps28bΔ (Ribosomal protein subunit-28B) mutants do not form PBs under normal growth conditions.Here, we demonstrate that the RPS28B 3'UTR is important for PB assembly, consistent with the fact that this is a known binding site for the PB assembly protein Edc3. However, expression of the RPS28B 3'UTR in isolation is insufficient to drive normal P-body assembly. Intriguingly, chimeric mRNA studies revealed that Rps28 protein, translated in cis from an mRNA bearing the RPS28B 3'UTR, physically interacts more strongly with Edc3 than Rps28 protein synthesized in trans. This Edc3-Rps28 interaction in turn also facilitates PB assembly. In summary, our work indicates that PB assembly may be preferentially nucleated by specific RNA "scaffolds", which may be a common theme in RNP granule assembly. Furthermore, this is the first description to our knowledge of assembly of a macromolecular complex being driven by a cis-translated protein interacting with another protein in the 3'UTR of the mRNA which encoded it.

3′ UTRs Regulate Protein Functions by Providing a Nurturing Niche during Protein Synthesis

“…In our hands, all so far tested 3 0 UTRs controlled protein activity without affecting protein abundance, indicating that this function of 3 0 UTRs is widespread. This has also been suggested by others (Fernandes 2019;Ribero 2019). 3' UTR-independent interactors 3' UTR-dependent interactors Figure 3.…”

Endogenous p53 expression in human and mouse is not regulated by its 3′UTR

“…Notably, a recent study that deleted 3′UTR sequences in several cytokine genes found similar discrepancies between reporter-based assays and gene expression from native contexts (Zhao et al, 2017). Although our data indicate that p53 abundance regulation is 3′UTR-independent, the 3′UTR may still have important functions possibly through control of protein localization or protein activity as has been shown for other proteins (Berkovits and Mayr, 2015; Moretti et al, 2015; Terenzio et al, 2018; Lee and Mayr, 2019; Fernandes and Buchan, 2020; Bae et al, 2020; Kwon, 2020; Mayr, 2019).…”

RNA Granules: A View from the RNA Perspective

“…In vitro, a small amount of RNA initially promotes protein droplet formation and afterwards begins dissolving the droplets once the concentration of the RNA reaches a threshold [ 76 , 77 ]. RNAs change physical properties of condensates (i.e., viscosity), augment granule assembly in vitro and in vivo [ 13 , 44 , 67 , 76 , 77 , 78 , 79 , 80 , 81 , 82 , 83 , 84 ], determine the three dimensional structure of granules in an RNA concentration-dependent manner [ 49 , 85 ], and can even instruct the morphology of granules by their ability to engage in intermolecular RNA-RNA interactions [ 86 ].…”