Alzheimer’s disease (AD) is characterized by overproduction of β amyloid peptides in the brain with progressive loss of neuronal cells. The 42-aa form of the β amyloid peptide (Aβ42) is implied as a major causative factor, because it is toxic to neurons and elicits inflammatory responses in the brain by activating microglial cells. Despite the overproduction of Aβ42, AD brain tissue also generates protective factor(s) that may antagonize the neurodestructive effect of Aβ42. Humanin is a gene cloned from an apparently normal region of an AD brain and encodes a 24-aa peptide. Both secreted and synthetic Humanin peptides protect neuronal cells from damage by Aβ42, and the effect of Humanin may involve putative cellular receptor(s). To elucidate the molecular identity of such receptor(s), we examined the activity of synthetic Humanin on various cells and found that Humanin induced chemotaxis of mononuclear phagocytes by using a human G protein-coupled formylpeptide receptor-like-1 (FPRL1) and its murine counterpart FPR2. Coincidentally, FPRL1 and FPR2 are also functional receptors used by Aβ42 to chemoattract and activate phagocytic cells. Humanin reduced the aggregation and fibrillary formation by suppressing the effect of Aβ42 on mononuclear phagocytes. In neuroblast cells, Humanin and Aβ42 both activated FPRL1; however, only Aβ42 caused apoptotic death of the cells, and its cytopathic effect was blocked by Humanin. We conclude that Humanin shares human FPRL1 and mouse FPR2 with Aβ42 and suggest that Humanin may exert its neuroprotective effects by competitively inhibiting the access of FPRL1 to Aβ42.

The E3 ubiquitin ligase Casitas B lymphoma protein (Cbl) controls the ubiquitin-dependent degradation of EGF receptor (EGFR), but its role in regulating downstream signaling elements with which it associates and its impact on biological outcomes of EGFR signaling are less clear. Here, we demonstrate that stimulation of EGFR on human mammary epithelial cells disrupts adherens junctions (AJs) through Vav2 and Rac1/Cdc42 activation. In EGF-stimulated cells, Cbl regulates the levels of phosphorylated Vav2 thereby attenuating Rac1/Cdc42 activity. Knockdown of Cbl and Cbl-b enhanced the EGF-induced disruption of AJs and cell motility. Overexpression of constitutively active Vav2 activated Rac1/Cdc42 and reorganized junctional actin cytoskeleton; these effects were suppressed by WT Cbl and enhanced by a ubiquitin ligase-deficient Cbl mutant. Cbl forms a complex with phospho-EGFR and phospho-Vav2 and facilitates phospho-Vav2 ubiquitinylation. Cbl can also interact with Vav2 directly in a Cbl Tyr-700-dependent manner. A ubiquitin ligase-deficient Cbl mutant enhanced the morphological transformation of mammary epithelial cells induced by constitutively active Vav2; this effect requires an intact Cbl Tyr-700. These results indicate that Cbl ubiquitin ligase plays a critical role in the maintenance of AJs and suppression of cell migration through down-regulation of EGFR-Vav2 signaling.

Sumoylation is one of the most essential mechanisms of reversible protein post-translational modifications and is a crucial biochemical process in the regulation of a variety of important biological functions. Sumoylation is also closely involved in various human diseases. The accurate computational identification of sumoylation sites in protein sequences aids in experimental design and mechanistic research in cellular biology. In this study, we introduced amino acid hydrophobicity as a parameter into a traditional binary encoding scheme and developed a novel sumoylation site prediction tool termed SUMOhydro. With the assistance of a support vector machine, the proposed method was trained and tested using a stringent non-redundant sumoylation dataset. In a leave-one-out cross-validation, the proposed method yielded an excellent performance with a correlation coefficient, specificity, sensitivity and accuracy equal to 0.690, 98.6%, 71.1% and 97.5%, respectively. In addition, SUMOhydro has been benchmarked against previously described predictors based on an independent dataset, thereby suggesting that the introduction of hydrophobicity as an additional parameter could assist in the prediction of sumoylation sites. Currently, SUMOhydro is freely accessible at http://protein.cau.edu.cn/others/SUMOhydro/.