β-PIX plays an important role in regulation of intestinal epithelial restitution by interacting with GIT1 and Rac1 after wounding

Abstract

Early gut mucosal restitution is a process by which intestinal epithelial cells (IECs) migrate over the wounded area, and its defective regulation occurs commonly in various critical pathological conditions. This rapid reepithelialization is mediated by different activating small GTP-binding proteins, but the exact mechanism underlying this process remains largely unknown. Recently, it has been reported that interaction between p21-activated kinase-interacting exchange factor (β-PIX) and G protein-coupled receptor kinase-interacting protein 1 (GIT1) activates small GTPases and plays an important role in the regulation of cell motility. Here, we show that induced association of β-PIX with GIT1 is essential for the stimulation of IEC migration after wounding by activating Rac1. Levels of β-PIX and GIT1 proteins and their association in differentiated IECs (line of IEC-Cdx2L1) were much higher than those observed in undifferentiated IECs (line of IEC-6), which was associated with an increase in IEC migration after wounding. Decreased levels of endogenous β-PIX by its gene-silencing destabilized β-PIX/GIT1 complexes, repressed Rac1 activity and inhibited cell migration over the wounded area. In contrast, ectopic overexpression of β-PIX increased the levels of β-PIX/GIT1 complexes, stimulated Rac1 activity, and enhanced intestinal epithelial restitution. Increased levels of cellular polyamines also stimulated β-PIX/GIT1 association, increased Rac1 activity, and promoted the epithelial restitution. Moreover, polyamine depletion decreased cellular abundances of β-PIX/GIT1 complex and repressed IEC migration after wounding, which was rescued by ectopic overexpression of β-PIX or GIT1. These results indicate that β-PIX/GIT1/Rac1 association is necessary for stimulation of IEC migration after wounding and that this signaling pathway is tightly regulated by cellular polyamines. NEW & NOTEWORTHY Our current study demonstrates that induced association of β-PIX with GIT1 is essential for the stimulation of intestinal epithelial restitution by activating Rac1, and this signaling pathway is tightly regulated by cellular polyamines.