UBE2O ubiquitinates PTRF/CAVIN1 and inhibits the secretion of exosome-related PTRF/CAVIN1.

BACKGROUND: Exosomes are small vesicles released by cells, which have crucial functions in intercellular communication. Exosomes originated from cell membrane invagination and are released followed by multivesicular bodies (MVBs) fused with the cell membrane. It is known that Polymerase I and Transcript Release Factor (PTRF, also known as Caveolin-associated Protein-1, CAVIN1) plays an important role in caveolae formation and exosome secretion. And PTRF in exosomes has been identified as a potential biomarker in multiple malignancies such as glioma and renal cell carcinoma. However, the mechanisms of how to regulate the secretion of exosome-related PTRF remain unknown. METHODS: We performed exogenous and endogenous immunoprecipitation assays to investigate the interaction between ubiquitin-conjugating enzyme E2O (UBE2O) and PTRF. We identified UBE2O ubiquitinated PTRF using ubiquitination assays. Then, exosomes were isolated by ultracentrifugation and identified by transmission electronic microscopy, western blot and nanoparticle tracking analysis. The effect of UBE2O on the secretion of exosome-related PTRF was analyzed by western blot, and the effect of UBE2O on exosome secretion was evaluated by exosome markers and the total protein content of exosomes. RESULTS: Here, we showed that UBE2O interacts with PTRF directly and ubiquitinates PTRF. Functionally, we found that UBE2O inhibited the effects of PTRF on exosome secretion via decreasing caveolae formation. Importantly, UBE2O decreased exosome secretion, resulting in downregulating PTRF secretion via exosomes. Our study also identified Serum Deprivation Protein Response (SDPR, also known as Caveolin-associated Protein-2, CAVIN2) interacted with both UBE2O and PTRF. Furthermore, we found that SDPR promotes PTRF expression in exosomes. Interestingly, even in the presence of SDPR, UBE2O still inhibited the secretion of exosome-related PTRF. CONCLUSIONS: Our study demonstrated that UBE2O downregulated exosome release and controlled the secretion of exosome-related PTRF through ubiquitinating PTRF. Since exosomes play an important role in malignant tumor growth and PTRF included in exosomes is a biomarker for several malignant tumors, increasing UBE2O expression in cells has the potential to be developed as a novel approach for cancer treatment. Video Abstract.

Imidazole-based pinanamine derivatives: Discovery of dual inhibitors of the wild-type and drug-resistant mutant of the influenza A virus.

We previously reported potent hit compound 4 inhibiting the wild-type influenza A virus A/HK/68 (H3N2) and A/M2-S31N mutant viruses A/WS/33 (H1N1), with its latter activity quite weak. To further increase its potency, a structure-activity relationship study of a series of imidazole-linked pinanamine derivatives was conducted by modifying the imidazole ring of this compound. Several compounds of this series inhibited the amantadine-sensitive virus at low micromolar concentrations. Among them, 33 was the most potent compound, which was identified as being active on an amantadine-sensitive virus through blocking of the viral M2 ion channel. Furthermore, 33 markedly inhibited the amantadine-resistant virus (IC50 = 3.4 µM) and its activity increased by almost 24-fold compared to initial compound, with its action mechanism being not M2 channel mediated.