Crystal Structure of Human Edc3 and Its Functional Implications

Abstract

Edc3 is an enhancer of decapping and serves as a scaffold that aggregates mRNA ribonucleoproteins together for P-body formation. Edc3 forms a network of interactions with the components of the mRNA decapping machinery and has a modular domain architecture consisting of an N-terminal Lsm domain, a central FDF domain, and a C-terminal YjeF-N domain. We have determined the crystal structure of the N-terminally truncated human Edc3 at a resolution of 2.2 Å. The structure reveals that the YjeF-N domain of Edc3 possesses a divergent Rossmann fold topology that forms a dimer, which is supported by sedimentation velocity and sedimentation equilibrium analysis in solution. The dimerization interface of Edc3 is highly conserved in eukaryotes despite the overall low sequence homology across species. Structure-based sitedirected mutagenesis revealed dimerization is required for efficient RNA binding, P-body formation, and likely for regulating the yeast Rps28B mRNA as well, suggesting that the dimeric form of Edc3 is a structural and functional unit in mRNA degradation.The controlled turnover of eukaryotic mRNA is crucial for regulating gene expression (11,25,29,42). Two general pathways exist to degrade eukaryotic mRNAs, both of which are initiated by deadenylation (23,35,39). Following deadenylation, mRNAs can be degraded 3Ј to 5Ј by the cytoplasmic exosome or, more commonly, are decapped by the Dcp1/Dcp2 decapping enzyme and degraded by the 5Ј-to-3Ј exonuclease, Xrn1p. The formation of an mRNA ribonucleoprotein (mRNP) capable of decapping correlates with the mRNA ceasing translation and forming a translationally repressed mRNA (13, 38). These translationally repressed mRNPs can aggregate in the cytoplasm to form processing bodies (P-bodies), which are dynamic cytoplasmic RNA granules. P-bodies are of interest, as the mRNPs within them have been implicated in translation repression (13, 26), general mRNA decay (14, 44), nonsense-mediated mRNA decay (45, 49), microRNA-mediated translational repression (32, 41), and mRNA storage (5, 6).An important aspect of understanding the process of mRNA decapping and P-body formation is to understand the interactions and functions of the proteins that form the translationally repressed mRNP capable of decapping and P-body localization. A variety of genetic, biochemical, cell biology, and genomic analyses have indicated that the mRNP capable of decapping and P-body localization contains a variety of conserved proteins that interact with each other and RNA, including the decapping enzyme Dcp1p/Dcp2p, the decapping activators Dhh1p, Lsm1-7p, Pat1p, and Edc3p, the 5Ј-3Ј exoribonuclease Xrn1p, and the Ccr4p-Pop2-Not1p deadenylase complex (20,38). It has also been shown that P-body formation can be affected by the pool of nontranslating mRNAs (20,38) and that proteins such as RCK/p54/Dhh1 and Pat1p contribute to P-body formation by increasing the pool of nontranslating mRNAs (13, 40). Recently, Dcp2p was found to contribute to P-body formation, presumably through its multiple interactions wi...

DCP1 forms asymmetric trimers to assemble into active mRNA decapping complexes in metazoa

“…Trimerization of DCP1 reveals an unexpected connectivity and complexity of the decapping network in multicellular eukaryotes, as both EDC3 and EDC4 are known or presumed to form homodimers (11,13,15,21). We find that DCP1a can interact with DCP2 and EDC4 independently of the interaction with EDC3 and DDX6/RCK.…”

Cytoplasmic mRNP granules at a glance

“…Overexpression studies suggest that some of these might play important roles in the assembly of mRNPs into PBs, although further studies are needed to verify this (Behm-Ansmant et al, 2006;Coller and Parker, 2005;Eulalio et al, 2009;Fenger-Grøn et al, 2005;Haas et al, 2010;Jinek et al, 2008;Yu et al, 2005). Other protein domains might also be important, as exemplified by the C-terminal homodimerization domain of the decapping enhancer Edc3, which is important for PB formation in S. cerevisiae (Decker et al, 2007;Ling et al, 2008). Important questions for future studies include determining why, as all evidence suggests, PB assembly complexes only assemble into PBs when they are associated with RNA and what prevents polysomeassociated mRNPs from assembling into PBs.…”

Similar Modes of Interaction Enable Trailer Hitch and EDC3 To Associate with DCP1 and Me31B in Distinct Protein Complexes

“…The FDF motifs of LSm13-15 are embedded in low-complexity regions rich in glycine and arginine (1,2). In contrast, in LSm16 (known as enhancer of decapping-3 and referred to as EDC3 hereafter) the FDF motif is followed by a conserved C-terminal YjeF_N domain that adopts a divergent Rossman fold similar to one in the N-terminal domain of bacterial YjeF (1,2,32). EDC3 (LSm16) is known to enhance bulk mRNA decapping in yeast and is required for the decapping-dependent regulation of RPS28B mRNA and YRA1 pre-mRNA (4,16,29).…”

“…In addition, EDC3 self-associates through its C-terminal YjeF_N domain (32,50). Similarly, the FDF motif and the YjeF_N domain of the Saccharomyces cerevisiae Edc3 protein mediate the interaction with the yeast Me31B ortholog (Dhh1p) and self-association, respectively, demonstrating that these interactions are conserved (13,21,32,33).LSm13 to LSm15 represent a group of orthologous proteins in different species. These include S. cerevisiae Scd6p (LSm13), D. melanogaster Trailer Hitch (Tral or LSm15), Caenorhabditis elegans CAR-1, and vertebrate RAP55 (LSm14) (1, 2).…”