USP32 regulates late endosomal transport and recycling through deubiquitylation of Rab7.

The endosomal system is a highly dynamic multifunctional organelle, whose complexity is regulated in part by reversible ubiquitylation. Despite the wide-ranging influence of ubiquitin in endosomal processes, relatively few enzymes utilizing ubiquitin have been described to control endosome integrity and function. Here we reveal the deubiquitylating enzyme (DUB) ubiquitin-specific protease 32 (USP32) as a powerful player in this context. Loss of USP32 inhibits late endosome (LE) transport and recycling of LE cargos, resulting in dispersion and swelling of the late compartment. Using SILAC-based ubiquitome profiling we identify the small GTPase Rab7-the logistical centerpiece of LE biology-as a substrate of USP32. Mechanistic studies reveal that LE transport effector RILP prefers ubiquitylation-deficient Rab7, while retromer-mediated LE recycling benefits from an intact cycle of Rab7 ubiquitylation. Collectively, our observations suggest that reversible ubiquitylation helps switch Rab7 between its various functions, thereby maintaining global spatiotemporal order in the endosomal system.

Alternative polyadenylation and its impact on cellular processes.

Impact of mRNA processing and/or modifications has long been associated with gene expression regulation. Accumulating evidence shows alternative polyadenylation (APA), as an mRNA related process, to emerge as a widespread mechanism in gene expression regulation. Through selecting alternate (proximal or distal) polyadenylation signals on the 3'-UTR of pre-mRNAs, APA generates multiple transcript isoforms which may even create proteomic diversity. Depending on the use of proximal or distal polyadenylation sites, 3'-UTR lengths can vary in a tightly controlled manner in a spatial and temporal mode. Therefore, APA and its deregulation with potential consequences are highly relevant to normal and disease states. In this review, in light of recent findings in the literature, mechanism and types of APA and roles of APA in biological processes (i.e. proliferation, development, differentiation, and transformation) are discussed.

Matrix metalloprotease 16 expression is downregulated by microRNA-146a in spontaneously differentiating Caco-2 cells.

Cellular differentiation in the gut is vital in maintaining the cellular and functional specialization of the epithelial layer. MicroRNAs (miRNAs) have recently emerged as one of the key players in orchestrating the differentiation process in the gut. Using the spontaneously differentiating Caco-2 cell line, we observed an increased expression of miR-146a but not miR-146b in the course of differentiation. Bioinformatic analyses revealed that the membrane type matrix metalloprotease 16 (MMP16, MT3-MMP) was a predicted target of miR-146a and a decrease in the mRNA and protein expression of MMP16 was observed in the course of differentiation. Transfection of a luciferase reporter vector containing the 3'UTR of MMP16 showed decreased luciferase activity due to miR-146a expression. With forced expression of miR-146a in undifferentiated Caco-2 cells, a decrease in the mRNA and protein levels of MMP16 and a lower gelatinase activity in a gelatin zymogram were observed. Additionally, forced expression of miR-146a in HT-29 colon cancer cells also resulted in decreased expression of MMP16, along with a decrease in the invasion through Matrigel. Taken together, we have shown here that MMP16 is regulated by miR-146a in spontaneously differentiated Caco-2 cells. As MMP16 activates the zymogen of MMP2, which is known to degrade extracellular matrix proteins, the regulation of MMP16 by miR-146a may account, at least in part, for lower motility of well-differentiated cells.