No abstract

Abstract Studies over the past three years have substantially expanded the involvements of eukaryotic initiation factor 3 (eIF3) in messenger RNA (mRNA) translation. It now appears that this multi-subunit complex is involved in every possible form of mRNA translation, controlling every step of protein synthesis from initiation to elongation, termination, and quality control in positive as well as negative fashion. Through the study of eIF3, we are beginning to appreciate protein synthesis as a highly integrated process coordinating protein production with protein folding, subcellular targeting, and degradation. At the same time, eIF3 subunits appear to have specific functions that probably vary between different tissues and individual cells. Considering the broad functions of eIF3 in protein homeostasis, it comes as little surprise that eIF3 is increasingly implicated in major human diseases and first attempts at therapeutically targeting eIF3 have been undertaken. Much remains to be learned, however, about subunit- and tissue-specific functions of eIF3 in protein synthesis and disease and their regulation by environmental conditions and post-translational modifications.

eIF3 is a multi-subunit complex with numerous functions in canonical translation initiation, including mRNA recruitment to the 40S ribosome, scanning for the start codon, and inhibition of premature 60S subunit joining 1-3 . eIF3 was also found to interact with 40S and 60S ribosomal proteins and translation elongation factors 4 , but a direct involvement in translation elongation has never been demonstrated. Using ribosome profiling, we found that eIF3 deficiency reduced early ribosomal elongation speed between codons 25 and 75 on a set of ~2,700 mRNAs encoding proteins associated with mitochondrial and membrane functions, resulting in defective synthesis of their encoded proteins. To promote early elongation, eIF3 forms stable protein interactions with 80S ribosomes translating the first ~60 codons and serves as a platform to recruit protein quality control factors, functions required for normal mitochondrial physiology. Accordingly, eIF3e +/knockout mice accumulate defective mitochondria in skeletal muscle and show a progressive decline in muscle strength with age. Hence, in addition to its canonical role in translation initiation, eIF3 interacts with 80S ribosomes to enhance, at the level of early elongation, the synthesis of proteins with membrane-associated functions, an activity that is critical for mitochondrial physiology and muscle health.The interaction of the 13 subunit eIF3 complex with mRNA is central to all forms of translation initiation, including cap-dependent and cap-independent mechanisms 5-7 . eIF3 promotes ternary complex recruitment, binding of the 43S pre-initiation complex to mRNA, as well as scanning processivity and fidelity 1,2 . Although eIF3 has traditionally been thought to be released from ribosomes upon 40S-60S subunit joining in vitro 8 , recent evidence in yeast 3 suggested that it can remain associated with 80S ribosomes during translation of short uORFs in order to facilitate re-initiation on downstream ORFs 9 . In addition, pull-down experiments indicated that yeast eIF3 forms stable in vivo interactions with factors involved in translation elongation (40S and 60S ribosomal proteins, eukaryotic elongation factors (eEFs), and tRNA synthetases), suggesting an unexplored role in integrating translation initiation with elongation 4 .

eIF3, whose subunits are frequently overexpressed in cancer, regulates mRNA translation from initiation to termination, but mRNA-selective functions of individual subunits remain poorly defined. Using multi-omic profiling upon acute depletion of eIF3 subunits, we observed that while eIF3a, b, e, and f markedly differed in their impact on eIF3 holo-complex formation and translation, they were each required for cancer cell proliferation and tumor growth. Remarkably, eIF3k showed the opposite pattern with depletion promoting global translation, cell proliferation, tumor growth, and stress resistance through repressing the synthesis of ribosomal proteins, especially RPS15A. Whereas ectopic expression of RPS15A mimicked the anabolic effects of eIF3k depletion, disruption of eIF3 binding to the 5'-UTR of RSP15A mRNA negated them. Supported by mathematical modeling, our data uncovers eIF3k-l as a mRNA-specific module which, through controlling RPS15A translation, serves as a rheostat of ribosome content to secure spare translational capacity that can be mobilized during stress.

Studies over the past three years have substantially expanded the involvements of eIF3 in mRNA translation. It now appears that this multi-subunit complex is involved in every possible form of mRNA translation, controlling every step of protein synthesis from initiation to elongation, termination and quality control in positive as well as negative fashion. Through the study of eIF3, we are beginning to appreciate protein synthesis as a highly integrated process coordinating protein production with protein folding, subcellular targeting, and degradation. At the same time, eIF3 subunits appear to have specific functions that probably vary between different tissues and individual cells. Considering the broad functions of eIF3 in protein homeostasis, it comes as little surprise that eIF3 is increasingly implicated in major human diseases and first attempts at therapeutically targeting eIF3 have been undertaken. Much remains to be learned, however, about subunit-and tissue-specific functions of eIF3 in protein synthesis and disease and their regulation by environmental conditions and posttranslational modifications.