Eukaryotic translation initiation factor 3 (eIF3) is a key regulator of translation initiation, but its in vivo assembly and molecular functions remain unclear. Here we show that eIF3 from Neurospora crassa is structurally and compositionally similar to human eIF3. N. crassa eIF3 forms a stable 12-subunit complex linked genetically and biochemically to the 13(th) subunit, eIF3j, which in humans modulates mRNA start codon selection. Based on N. crassa genetic analysis, most subunits in eIF3 are essential. Subunits that can be deleted (e, h, k and l) map to the right side of the eIF3 complex, suggesting that they may coordinately regulate eIF3 function. Consistent with this model, subunits eIF3k and eIF3l are incorporated into the eIF3 complex as a pair, and their insertion depends on the presence of subunit eIF3h, a key regulator of vertebrate development. Comparisons to other eIF3 complexes suggest that eIF3 assembles around an eIF3a and eIF3c dimer, which may explain the coordinated regulation of human eIF3 levels. Taken together, these results show that Neurospora crassa eIF3 provides a tractable system for probing the structure and function of human-like eIF3 in the context of living cells.

Viral mRNAs that lack a 5' m7GTP cap and a 3' poly-A tail rely on structural elements in their untranslated regions (UTRs) to form unique RNA-protein complexes that regulate viral translation. Recent studies of the barley yellow dwarf virus (BYDV) have revealed eukaryotic initiation factor 3 (eIF3) plays a significant role in facilitating communication between its 5' and 3' UTRs by binding both UTRs simultaneously. This report uses in vitro translation assays, fluorescence anisotropy binding assays, and selective 2'-hydroxyl acylation analyzed by primer extension (SHAPE) footprinting to identify secondary structures that are selectively interacting with eIF3. SHAPE data also show that eIF3 alters its interaction with BYDV structures when another factor crucial for BYDV translation, eIF4F, is introduced by the 3' BYDV translational enhancer (BTE). The observed BTE and eIF4F-induced shift of eIF3 position on the 5' UTR and the translational effects of altering eIF3-binding structures (SLC and SLII) support a new model for BYDV translation initiation that requires the reorientation of eIF3 on BYDV UTRs. This eIF3 function in BYDV translation initiation is both reminiscent of and distinct from eIF3-RNA interactions found in other non-canonically translating mRNAs (e.g. HCV). This characterization of a new role in translation initiation expands the known functionality of eIF3 and may be broadly applicable to other non-canonically translating mRNAs.

To describe the prevalence and clinical associations of autoantibodies to a novel autoantigen, eukaryotic initiation factor 3 (eIF3), detected in idiopathic inflammatory myositis.

The eukaryotic translation initiation factor 3 (eIF3) has multiple roles during the initiation of translation of cytoplasmic mRNAs. How individual subunits of eIF3 contribute to the translation of specific mRNAs remains poorly understood, however. This is true in particular for those subunits that are not conserved in budding yeast, such as eIF3h.

Malaria drug resistance is hampering the fight against the deadliest parasitic disease affecting over 200 million people worldwide. We recently developed quinoline-quinazoline-based inhibitors (as compound 70) as promising new antimalarials. Here, we aimed to investigate their mode of action by using thermal proteome profiling (TPP). The eukaryotic translation initiation factor 3 (EIF3i) subunit I was identified as the main target protein stabilized by compound 70 in Plasmodium falciparum. This protein has never been characterized in malaria parasites. P. falciparum parasite lines were generated expressing either a HA tag or an inducible knockdown of the PfEIF3i gene to further characterize the target protein. PfEIF3i was stabilized in the presence of compound 70 in a cellular thermal shift Western blot assay, pointing that PfEIF3i indeed interacts with quinoline-quinazoline-based inhibitors. In addition, PfEIF3i-inducible knockdown blocks intra-erythrocytic development in the trophozoite stage, indicating that it has a vital function. We show that PfEIF3i is mostly expressed in late intra-erythrocytic stages and localizes in the cytoplasm. Previous mass spectrometry reports show that PfEIF3i is expressed in all parasite life cycle stages. Further studies will explore the potential of PfEIF3i as a target for the design of new antimalarial drugs active all along the life cycle of the parasite.

The main role of the translation initiation factor 3 (eIF3) is to orchestrate formation of 43S-48S preinitiation complexes (PICs). Until now, most of our knowledge on eIF3 functional contribution to regulation of gene expression comes from yeast studies. Hence, here we developed several novel in vivo assays to monitor the integrity of the 13-subunit human eIF3 complex, defects in assembly of 43S PICs, efficiency of mRNA recruitment, and postassembly events such as AUG recognition. We knocked down expression of the PCI domain-containing eIF3c and eIF3a subunits and of eIF3j in human HeLa and HEK293 cells and analyzed the functional consequences. Whereas eIF3j downregulation had barely any effect and eIF3a knockdown disintegrated the entire eIF3 complex, eIF3c knockdown produced a separate assembly of the a, b, g, and i subunits (closely resembling the yeast evolutionary conserved eIF3 core), which preserved relatively high 40S binding affinity and an ability to promote mRNA recruitment to 40S subunits and displayed defects in AUG recognition. Both eIF3c and eIF3a knockdowns also severely reduced protein but not mRNA levels of many other eIF3 subunits and indeed shut off translation. We propose that eIF3a and eIF3c control abundance and assembly of the entire eIF3 and thus represent its crucial scaffolding elements critically required for formation of PICs.

Collybistin (CB), a neuron-specific guanine nucleotide exchange factor, has been implicated in targeting gephyrin-GABAA receptors clusters to inhibitory postsynaptic sites. However, little is known about additional CB partners and functions.

Microgametogenesis is the post-meiotic pollen developmental phase when unicellular microspores develop into mature tricellular pollen. In rice, microgametogenesis can influence grain yields to a great degree because pollen abortion occurs more easily during microgametogenesis than during other stages of pollen development. However, our knowledge of the genes involved in microgametogenesis in rice remains limited. Due to the dependence of pollen development on the regulatory mechanisms of protein expression, we identified the encoding gene of the eukaryotic translation initiation factor 3, subunit f in Oryza sativa (OseIF3f). Immunoprecipitation combined with mass spectrometry confirmed that OseIF3f was a subunit of rice eIF3, which consisted of at least 12 subunits including eIF3a, eIF3b, eIF3c, eIF3d, eIF3e, eIF3f, eIF3g, eIF3h, eIF3i, eIF3k, eIF3l, and eIF3m. OseIF3f showed high mRNA levels in immature florets and is highly abundant in developing anthers. Subcellular localization analysis showed that OseIF3f was localized to the cytosol and the endoplasmic reticulum in rice root cells. We further analyzed the biological function of OseIF3f using the double-stranded RNA-mediated interference (RNAi) approach. The OseIF3f-RNAi lines grew normally at the vegetative stage but displayed a large reduction in seed production and pollen viability, which is associated with the down-regulation of OseIF3f. Further cytological observations of pollen development revealed that the OseIF3f-RNAi lines showed no obvious abnormalities at the male meiotic stage and the unicellular microspore stage. However, compared to the wild-type, OseIF3f-RNAi lines contained a higher percentage of arrested unicellular pollen at the bicellular stage and a higher percentage of arrested unicellular and bicellular pollen, and aborted pollen at the tricellular stage. These results indicate that OseIF3f plays a role in microgametogenesis.

Prevention of genital herpes is a global health priority. B5, a recently identified ubiquitous human protein, was proposed as a candidate HSV entry receptor. The current studies explored its role in HSV infection. Viral plaque formation was reduced by approximately 90% in human cells transfected with small interfering RNA targeting B5 or nectin-1, an established entry receptor. However, the mechanisms were distinct. Silencing of nectin-1 prevented intracellular delivery of viral capsids, nuclear transport of a viral tegument protein, and release of calcium stores required for entry. In contrast, B5 silencing had no effect on these markers of entry, but inhibited viral protein translation. Specifically, viral immediate early genes, ICP0 and ICP4, were transcribed, polyadenylated and transported from the nucleus to the cytoplasm, but the viral transcripts did not associate with ribosomes or polysomes in B5-silenced cells. In contrast, immediate early gene viral transcripts were detected in polysome fractions isolated from control cells. These findings are consistent with sequencing studies demonstrating that B5 is eukaryotic initiation factor 3 subunit m (eIF3m). Although B5 silencing altered the polysome profile of cells, silencing had little effect on cellular RNA or protein expression and was not cytotoxic, suggesting that this subunit is not essential for host cellular protein synthesis. Together these results demonstrate that B5 plays a major role in the initiation of HSV protein translation and could provide a novel target for strategies to prevent primary and recurrent herpetic disease.

Head and neck squamous cell carcinoma (HNSCC) is the sixth most common cancer type worldwide. Deregulation of mRNA translation is a frequent feature of cancer. Eukaryotic translation initiation factor 3 subunit B (EIF3B) has been reported as an oncogene; however, its role in HNSCC has yet to be fully elucidated.

Liver cancer is one of the major malignancies with the worst prognosis among all solid tumor types. It is therefore ponderable to explore prognostic biomarkers and therapeutic targets for liver cancer. Eukaryotic translation initiation factor 3 subunit B (EIF3B) is closely linked to the transcription initiation of cancer-associated genes. In the present study, EIF3B was indicated to be a potential prognostic biomarker of liver cancer. The mRNA expression level of EIF3B in liver cancer was assessed by analyzing the Cancer Genome Atlas dataset. χ2 and Fisher's exact tests were used to assess the association of EIF3B expression with clinical parameters. Receiver-operating characteristic curve analysis was used for evaluating the diagnostic value of EIF3B. Overall and relapse-free survival were assessed using Kaplan-Meier curves to determine the association between EIF3B expression and survival. Univariate and multivariate Cox regression analysis were performed to identify the factors affecting overall/relapse-free survival. Gene set enrichment analysis (GSEA) was used to identify signaling pathways associated with EIF3B in liver cancer. It was revealed that EIF3B was highly expressed in liver cancer tissues and it had a promising diagnostic ability. Furthermore, the survival analysis indicated that patients with high EIF3B expression generally had shorter overall as well as relapse-free survival. Univariate and multivariate Cox analysis suggested that high EIF3B mRNA expression may serve as an independent biomarker for the prognostication of patients with liver cancer. GSEA suggested that MYC-V1 (HALLMARK_MYC_TARGETS_V1 geneset; P=0.009), MYC-V2 (HALLMARK_MYC_TARGETS_V2 geneset; P=0.004) and DNA repair pathways (HALLMARK_DNA_REPAIR geneset; P<0.001) were differentially enriched in high EIF3B expression and low EIF3B expression groups. In conclusion, high EIF3B expression was indicated to be an independent prognostic biomarker for patients with liver cancer.

Eukaryotic translation initiation factor 3 (eIF3) is a large protein complex that participates in most translation initiation processes. While eIF3 has been well characterized, less is known about the roles of individual eIF3 subunits, particularly in plants. Here, we identified and characterized OseIF3e in rice (Oryza sativa L.). OseIF3e was constitutively expressed in various tissues, but most strongly in vigorously growing organs. Transgenic OseIF3e-silenced rice plants showed inhibited growth in seedling and vegetative stages. Repression of OseIF3e led to defects in pollen maturation but did not affect pollen mitosis. In rice, eIF3e interacted with eIF3 subunits b, d, e, f, h, and k, and with eIF6, forming homo- and heterodimers to initiate translation. Furthermore, OseIF3e was shown by yeast two-hybrid assay to specifically bind to inhibitors of cyclin-dependent kinases 1, 5, and 6. This interaction was mediated by the sequence of amino acid residues at positions 118-138, which included a conserved motif (IGPEQIETLYQFAKF). These results suggested although OseIF3e is not a "functional core" subunit of eIF3, it still plays crucial roles in rice growth and development, in combination with other factors. We proposed a pathway by which OseIF3e influence organ size and pollen maturation in rice, providing an opportunity to optimize plant architecture for crop breeding.

Mammalian eukaryotic translation initiation factor 3 (eIF3) is the largest complex of the translation initiation factors. The eIF3 complex is comprised of thirteen subunits, which are named eIF3a to eIF3 m in most multicellular organisms. The eIF3e gene locus is one of the most frequent integration sites of mouse mammary tumor virus (MMTV), which induces mammary tumors in mice. MMTV-integration events result in the expression of C-terminal-truncated eIF3e proteins, leading to mammary tumor formation. We have shown that tumor formation can be partly caused by activation of hypoxia-inducible factor 2α. To investigate the function of eIF3e in mammals, we generated eIF3e-deficient mice. These eIF3e-/- mice are embryonically lethal, while eIF3e+/- mice are much smaller than wild-type mice. In addition, eIF3e+/- mouse embryonic fibroblasts (MEFs) contained reduced levels of eIF3a and eIF3c subunits and exhibited reduced cellular proliferation. These results suggest that eIF3e is essential for embryonic development in mice and plays a role in maintaining eIF3 integrity.

Previous studies have shown that subunits E (eIF3e), F (eIF3f) and H (elF3h) of eukaryotic translation initiation factor 3 play important roles in cell development in humans and yeast. eIF3e and eIF3h have also been reported to be important for normal cell growth in Arabidopsis. However, the functions of subunit eIF3f remain largely unknown in plant species. Here we report characterization of mutants for the Arabidopsis eIF3f (AteIF3f) gene. AteIF3f encodes a protein that is highly expressed in pollen grains, developing embryos and root tips, and interacts with Arabidopsis eIF3e and eIF3h proteins. A Ds insertional mutation in AteIF3f disrupted pollen germination and embryo development. Expression of some of the genes that are essential for pollen tube growth and embryogenesis is down-regulated in ateif3f-1 homozygous seedlings obtained by pollen rescue. These results suggested that AteIF3f might play important roles in Arabidopsis cell growth and differentiation in combination with eIF3e and eIF3h.

Inappropriate surface expression of voltage-gated Ca(2+)channels (CaV) in pancreatic ß-cells may contribute to the development of type 2 diabetes. First, failure to increase intracellular Ca(2+) concentrations at the sites of exocytosis impedes insulin release. Furthermore, excessive Ca(2+) influx may trigger cytotoxic effects. The regulation of surface expression of CaV channels in the pancreatic β-cells remains unknown. Here, we used real-time 3D confocal and TIRFM imaging, immunocytochemistry, cellular fractionation, immunoprecipitation and electrophysiology to study trafficking of L-type CaV1.2 channels upon β-cell stimulation. We found decreased surface expression of CaV1.2 and a corresponding reduction in L-type whole-cell Ca(2+) currents in insulin-secreting INS-1 832/13 cells upon protracted (15-30 min) stimulation. This internalization occurs by clathrin-dependent endocytosis and could be prevented by microtubule or dynamin inhibitors. eIF3e (Eukaryotic translation initiation factor 3 subunit E) is part of the protein translation initiation complex, but its effect on translation are modest and effects in ion channel trafficking have been suggested. The factor interacted with CaV1.2 and regulated CaV1.2 traffic bidirectionally. eIF3e silencing impaired CaV1.2 internalization, which resulted in an increased intracellular Ca(2+) load upon stimulation. These findings provide a mechanism for regulation of L-type CaV channel surface expression with consequences for β-cell calcium homeostasis, which will affect pancreatic β-cell function and insulin production.

The EIF3 gene family is essential in controlling translation initiation during the cell cycle. The significance of the EIF3 subunits as prognostic markers and therapeutic targets in breast cancer is not yet clear. We analyzed the expression of EIF3 subunits in breast cancer on the GEPIA and Oncomine databases and compared their expression in breast cancer and normal tissues using BRCA data downloaded from TCGA. Then we performed clinical survival analysis on the Kaplan-Meier Plotter database and clinicopathologic analysis on the bc-genexMiner v4.1 database. And EIF3B was chosen for mutation analysis via the Cancer SEA online tool. Meanwhile, we performed the immunohistochemical assay, real-time RT-PCR, and Western blotting to analyze EIF3B expression levels in breast cancer. An EIF3B knockdown and a negative control cell line were conducted for MTT assay and cell cycle analysis to assess cell growth. Specifically, the results of TCGA and online databases demonstrated that upregulated EIF3B was associated with poorer overall and advanced tumor progression. We also confirmed that EIF3B was more highly expressed in breast cancer cells and tissues than normal and correlated with a worse outcome. And knockdown of EIF3B expression inhibited the cell cycle and proliferation. Furthermore, EIF3B was highly mutated in breast cancer. Collectively, our results suggested EIF3B as a potential prognostic marker and therapeutic target for breast cancer.

Background: Pancreatic cancer (PC) is a malignant tumor with hidden incidence, high degree of malignancy, rapid disease progression, and poor prognosis. Eukaryotic translation initiation factor 3 subunit B (EIF3B) is necessary for tumor growth, which is an alternative therapeutic target for many cancers. However, little is known about the relationship between EIF3B and PC. Methods: The expression of EIF3B in PC was detected by immunohistochemistry. EIF3B knockdown cell models were constructed by lentivirus infection. The MTT assay, the wound-healing assay, the transwell assay, the flow cytometry, and the Human Apoptosis Antibody Array was used to detect the effects of EIF3B knockdown on cell proliferation, cell migration, cell apoptosis, and cell cycle in vitro. Also, the effects of EIF3B knockdown on the tumor growth of PC were determined in vivo. Results: This study showed that the expression level of EIF3B was significantly up-regulated in PC tumor tissues and associated with pathological grade. In vitro, EIF3B knockdown inhibited the PC cell proliferation and migration, and the apoptosis levels were obviously promoted by regulating apoptosis-related proteins including Bcl-2, HSP27, HSP60, Survivin, sTNF-R2, TNF-α, TNF-β, TRAILR-3, TRAILR-4, and XIAP. Furthermore, the tumor growth of PC was inhibited after the knockdown of EIF3B in vivo. Conclusion: EIF3B was up-regulated in PC and was a promoter in the development and progression of PC, which could be considered as a therapeutic target for the treatment of PC.

The gene encoding eIF3g (TaeIF3g), one of the 11 subunits of eukaryotic translation initiation factor 3 (eIF3), was cloned from wheat for carrying out its functional analysis. Transgenic expression of TaeIF3g enhanced the tolerance of TaeIF3g-overexpressing parental yeast cells and Arabidopsis plants under different abiotic stress conditions. Compared to untransformed plants, TaeIF3g-overexpressing Arabidopsis thaliana plants exhibited significantly higher survival rate, soluble proteins and photosynthetic efficiency, and enhanced protection against photooxidative stress under drought conditions. This study provides first evidence that TaeIF3g imparts stress tolerance and could be a potential candidate gene for developing crop plants tolerant to abiotic stress.

Ovarian cancer is the most common cause of gynaecological cancer-associated death; thus, promising biomarkers and new therapeutic targets for ovarian cancer must be explored. Here, we report that eukaryotic translation initiation factor 3 subunit D (EIF3D), a member of the EIF3 family, was overexpressed in ovarian cancer clinical tissues. Furthermore, the expression of EIF3D was correlated with the International Federation of Gynecology and Obstetrics stage and pathological differentiation stage. 3-(4,5-dimethylthylthiazol-2-yl)-2 (MTT) and colony formation assays revealed that the lentivirus-mediated knockdown of EIF3D suppresses cell proliferation in the ovarian tumour cell lines CAOV-3 and SKOV-3. Flow cytometry revealed that cells were arrested at the G2/M phase of the cell cycle and that cyclin-dependent kinase 1 was also altered after EIF3D silencing. The results presented here demonstrate that EIF3D may play an important role in the occurrence and development of ovarian cancer.

Ovarian cancer remains the leading cause of death among all gynaecological cancers, illustrating the urgent need to understand the molecular mechanisms involved in this disease. Eukaryotic initiation factor 3c (EIF3c) plays an important role in protein translation and cancer cell growth and proliferation, but its role in human ovarian cancer is unclear. Our results showed that EIF3c silencing significantly up-regulated 217 and down-regulated 340 genes. Ingenuity Pathway Analysis (IPA) indicated that the top differentially expressed genes are involved in 'Classical Pathways', 'Diseases and Functions' and 'Networks', especially those involved in signalling and cellular growth and proliferation. In addition, eIF3c silencing inhibited cellular proliferation, enhanced apoptosis and regulated the expression of apoptosis-associated proteins. In conclusion, these results indicate that by dysregulating translational initiation, eIF3c plays an important role in the proliferation and survival of human ovarian cancer cells. These results should provide experimental directions for further in-depth studies on important human ovarian cancer cell pathways.