The prognostic impacts of PABPC1 expression on gastric cancer patients.

Aim: To assess the prognostic impacts of PABPC1 on gastric cancer (GC) patients. Methods: The expression levels of PABPC1 in GC tissues and normal gastric tissues were initially compared via bioinformatics analysis. Immunohistochemical staining was accomplished to assess the expression of PABPC1 in the included GC patients. Then the impacts of PABPC1 expression on survival of GC patients were evaluated by Cox regression and Kaplan-Meier analyses. Results: The expression levels of PABPC1 in gastric tissues were significantly higher than those in normal gastric tissues (paired, p = 0.002; unpaired, p = 3.60e-9). By Kaplan-Meier, it was demonstrated that high expression of PABPC1 was significantly associated with worse overall and disease-free survival. Furthermore, high PABPC1 expression was demonstrated to be an independent predictive factor for both overall (p = 0.013; hazard ratio = 2.058; 95% CI: 1.162-3.644) and disease-free (p = 0.018; hazard ratio = 2.284; 95% CI: 1.153-4.524) survival. Conclusion: PABPC1 is a potential prognostic biomarker for GC patients.

Lay abstract Previous studies have reported that PABPC1 is involved in a series of biological processes and participates in many cancers. However, the specific role of PABPC1 in different cancers varies significantly, and PABPC1 has not been fully investigated in gastric cancer (GC). In the present study, it was demonstrated that PABPC1 was significantly upregulated in GC and its high expression in GC was significantly associated with worse overall and disease-free survival, indicating the potential of PABPC1 as a novel prognostic biomarker for GC.

Distinct recruitment of human eIF4E isoforms to processing bodies and stress granules.

BACKGROUND: Eukaryotic translation initiation factor 4E (eIF4E) plays a pivotal role in the control of cap-dependent translation initiation, modulates the fate of specific mRNAs, occurs in processing bodies (PBs) and is required for formation of stress granules (SGs). In this study, we focused on the subcellular localization of a representative compendium of eIF4E protein isoforms, particularly on the less studied members of the human eIF4E protein family, eIF4E2 and eIF4E3. RESULTS: We showed that unlike eIF4E1, its less studied isoform eIF4E3_A, encoded by human chromosome 3, localized to stress granules but not PBs upon both heat shock and arsenite stress. Furthermore, we found that eIF4E3_A interacts with human translation initiation factors eIF4G1, eIF4G3 and PABP1 in vivo and sediments into the same fractions as canonical eIF4E1 during polysome analysis in sucrose gradients. Contrary to this finding, the truncated human eIF4E3 isoform, eIF4E3_B, showed no localization to SGs and no binding to eIF4G. We also highlighted that eIF4E2 may exhibit distinct functions under different stresses as it readily localizes to P-bodies during arsenite and heat stresses, whereas it is redirected to stress granules only upon heat shock. We extended our study to a number of protein variants, arising from alternative mRNA splicing, of each of the three eIF4E isoforms. Our results surprisingly uncovered differences in the ability of eIF4E1_1 and eIF4E1_3 to form stress granules in response to cellular stresses. CONCLUSION: Our comparison of all three human eIF4E isoforms and their protein variants enriches the intriguing spectrum of roles attributed to the eukaryotic initiation translation factors of the 4E family, which exhibit a distinctive localization within different RNA granules under different stresses. The localization of eIF4E3_A to stress granules, but not to processing bodies, along with its binding to eIF4G and PABP1 suggests a role of human eIF4E3_A in translation initiation rather than its involvement in a translational repression and mRNA decay and turnover. The localization of eIF4E2 to stress granules under heat shock but not arsenite stress indicates its distinct function in cellular response to these stresses and points to the variable protein content of SGs as a consequence of different stress insults.

Exosome-mediated extracellular release of polyadenylate-binding protein 1 in human metastatic duodenal cancer cells.

Exosomes are small vesicles secreted from cells that transport their embedded molecules through bidirectional exocytosis- and endocytosis-like pathways. Expression patterns of exosomal molecules such as proteins and RNAs can be indicative of cell type since their signature is thought to be unique among cells. Using human primary (AZ-521) and metastatic (AZ-P7a) duodenal cancer cell lines, we conducted a comparative exosomal proteome analysis to identify proteins with metastatic marker potential. As determined by LC-MS/MS and Western blot analyses, polyadenylate-binding protein 1 (PABP1) was found to be predominantly abundant in AZ-P7a exosomes. The amount of exosomal PABP1 in AZ-P7a cells increased by treating the cells with inhibitors for the classical ER/Golgi secretory pathway (brefeldin A and monensin) and the ubiquitin-proteasome pathway (MG-132 and PYR-41). Treatment of AZ-P7a cells with the neutral sphingomyelinase inhibitor GW4869, which suppresses exosome release, not only reduced the amount of exosomal PABP1 but also produced PABP1-immunoreactive products cleaved via a proteolysis-like process. Taken together, these results suggest that AZ-P7a cells do not tolerate intracellular PABP1 accumulation and are thus exported into the extracellular milieu by the exosome-mediated pathway. In addition, PABP1 has a potential use as a biomarker for metastatic duodenal cancer.

Induction of HSP70 expression and recruitment of HSC70 and HSP70 in the nucleus reduce aggregation of a polyalanine expansion mutant of PABPN1 in HeLa cells.

Nuclear inclusions formed by the aggregation of a polyalanine expansion mutant of the nuclear poly(A)-binding protein (PABPN1) is a hallmark of oculopharyngeal muscular dystrophy (OPMD). OPMD is a dominant autosomal disease in which patients exhibit progressive difficulty of swallowing and eyelid elevation, starting around the age of 50. At present, there is no specific treatment to reduce the aggregate burden in patients. However, in cell culture models of OPMD, reduction of protein aggregation can be achieved by ectopic expression of HSP70. As gene transfer may not be the most effective means to elevate HSP70 levels, we tested four pharmacological agents for their ability to induce HSP70, recruit both HSP70 and HSC70 into the cell nucleus and reduce mutant PABPN1 aggregation in a HeLa cell culture model. We show here that exposure to moderate levels of ZnSO4, 8-hydroxyquinoline, ibuprofen and indomethacin produced a robust stress response resulting in the induction of HSP70 in HeLa cells expressing the mutant PABPN1 as a green fluorescent protein (GFP) fusion protein. Both HSP70 and the constitutive chaperone HSC70 localized in the nucleus of cells treated with any one of the four agents. This stress response was similar to what was observed following hyperthermia. All four agents also caused a significant reduction in the cellular burden of protein aggregates, as was judged by confocal microscopy and solubility changes of the aggregates. A concomitant reduction of cell death in drug-treated mutant PABPN1 expressing cells was also observed.

Interaction of paxillin with poly(A)-binding protein 1 and its role in focal adhesion turnover and cell migration.

We have previously identified poly(A)-binding protein 1 (PABP1) as a ligand for paxillin and shown that the paxillin-PABP1 complex undergoes nucleocytoplasmic shuttling. By targeting the paxillin-binding subdomain sequences in PABP1, we have generated mutants of PABP1 that do not bind to cellular paxillin. Here we report that paxillin association is necessary for efficient nuclear export of PABP1 and that RNA interference of paxillin drives the nuclear accumulation of PABP1. Furthermore, ablation of paxillin-PABP1 association impeded a number of indices of cell motility including spreading on fibronectin, cell migration on two-dimensional matrices, and transmigration in Boyden chambers. These data indicate that PABP1 must associate with paxillin in order to be efficiently transported from the nucleus to the cytoplasm and that this event is necessary for cells to remodel their focal adhesions during cell migration.