Decapping and Decay of Messenger RNA Occur in Cytoplasmic Processing Bodies

Abstract

A major pathway of eukaryotic messenger RNA (mRNA) turnover begins with deadenylation, followed by decapping and 5′ to 3′ exonucleolytic decay. We provide evidence that mRNA decapping and 5′ to 3′ degradation occur in discrete cytoplasmic foci in yeast, which we call processing bodies (P bodies). First, proteins that activate or catalyze decapping are concentrated in P bodies. Second, inhibiting mRNA turnover before decapping leads to loss of P bodies; however, inhibiting turnover at, or after, decapping, increases the abundance and size of P bodies. Finally, mRNA degradation intermediates are localized to P bodies. These results define the flux of mRNAs between polysomes and P bodies as a critical aspect of cytoplasmic mRNA metabolism and a possible site for regulation of mRNA degradation.Messenger RNA turnover is an important step in regulating gene expression. In yeast, the major pathway of mRNA decay is initiated by shortening of the 3′ poly(adenosine) [poly(A)] tail, followed by removal of the cap by the decapping enzyme Dcp1p/Dcp2p, which in turn allows 5′ to 3′ exonucleolytic decay (1-8). Decapping is a key step in this pathway, because it permits the destruction of the mRNA and is a site of numerous control inputs (9).Several observations suggest that decapping occurs when the mRNA undergoes a transition from a translationally competent messenger ribonucleoprotein (mRNP) to an mRNP state destined for decay. For example, the translation initiation factor eIF4E, which binds the cap structure, is an inhibitor of decapping both in vitro and in vivo (10,11). Moreover, deadenylated mRNAs that interact with a complex of Lsm1-7 proteins, which activates decapping, are no longer bound by eIF4E or eIF4G (12).The hypothesis that mRNAs enter a non-translating state after deadenylation and before decapping is analogous to the storage of mRNA in numerous biological contexts where deadenylated mRNAs are translationally repressed before their later activation. Consistent with a mechanistic similarity between decapping and mRNA storage, Dhh1p, which is an activator of decapping in yeast (13), has homologs that are required for the translational repression and storage of maternal mRNAs in Xenopus, Drosophila, and Caenorhabditis (14-16). Such stored mRNAs are often localized in discrete cytoplasmic granules, which represent accumulations of translationally repressed mRNAs (15). This analogy suggested the possibility that Dhh1p

Overexpression of a LAM Domain Containing RNA-Binding Protein LARPIc Induces Precocious Leaf Senescence in Arabidopsis

“…In C. elegans, CeLARP1 was reported to be a cytoplasmic protein that localizes to processing bodies (P-bodies) (Nykamp et al, 2008), which are sites of mRNA decay and storage. Interestingly, we found that 1c-GFP foci were sensitive to cycloheximide treatment, an inhibitor that can disrupt P-body formation (Goeres et al, 2007;Sheth and Parker, 2003) (data not shown). Does LARP1c localize to P-bodies?…”

Small Interfering RNA-mediated Silencing Induces Target-dependent Assembly of GW/P Bodies

“…The majority of GWB disassembled before mitosis and small GWB reassembled in early G1 (Yang et al, 2004). Second, as sites for 5Ј33Ј mRNA decay, the size and number of GWB are affected by blocking deadenylation, decapping, and 5Ј33Ј mRNA degradation or translation (Sheth and Parker, 2003;Cougot et al, 2004;Andrei et al, 2005;Teixeira et al, 2005). GWB require RNA for assembly, and the amount of mRNA or mRNA decay intermediates accumulated in GWB affects the size and number of these foci (Sen and Blau, 2005;Brengues et al, 2005;Teixeira et al, 2005).…”

Principles and effects of microRNA-mediated post-transcriptional gene regulation

“…In cultured human cell miRNA-binding caused the targeted mRNA to localize to P-bodies, which is argued to occurs in response to the primary regulatory event, repression of translational initiation (Pillai et al, 2005;Bhattacharyya et al, 2006). Indeed, eIF4e and eIF4e-transporter proteins participate in the transport of mRNAs into the PBs (Andrei et al, 2005) and in yeast inhibition of translation initiation increases localization of an mRNA to PBs (Sheth and Parker, 2003).…”