Snail family proteins regulate transcription of molecules for cell-cell adhesion during epithelial-mesenchymal transition (EMT). Based on putative glycogen synthase kinase 3β (GSK-3β) phosphorylation sites within the Slug/Snail2, we explored the significance of GSK-3β-mediated phosphorylation in Slug/Snail2 expression during EMT. Mutation of the putative GSK-3β phosphorylation sites (S92/96A or S100/104A) enhanced the Slug/Snail2-mediated EMT properties of E-cadherin repression and vimentin induction, compared with wild-type Slug/Snail2. S92/96A mutation inhibited degradation of Slug/Snail2 and S100/104A mutation extended nuclear stabilization. Inhibition of GSK-3β activity caused similar effects, as did the phosphorylation mutations. Thus, our study suggests that GSK-3β-mediated phosphorylation of Slug/Snail2 controls its turnover and localization during EMT.

Actin remodeling plays a crucial role in insulin-induced translocation of glucose transporter 4 (GLUT4) from the cytoplasm to the plasma membrane and subsequent glucose transport. Protein kinase C (PKC) zeta has been implicated in this translocation process, although the exact mechanism remains unknown. In this study, we investigated the effect of PKCzeta on actin cytoskeleton and translocation of GLUT4 in CHO-K1 cells expressing myc-tagged GLUT4. Insulin stimulated the phosphorylation of PKCzeta at Thr410 with no apparent effect on its protein expression. Moreover, insulin promoted colocalization of PKCzeta and actin that could be abolished by Latrunculin B. The overexpression of PKCzeta mimicked the insulin-induced change in actin cytoskeleton and translocation of GLUT4. These effects were also completely abrogated by Latrunculin B treatment. Using cell-permeable pseudosubstrate (PS) inhibitor of PKCzeta, the response to insulin could be alleviated. Our results strongly suggest that PKCzeta mediates the stimulatory effect of insulin on GLUT4 translocation through its interaction with actin cytoskeleton.

This study constitutes the first report revealing the participation of prominin-1 in glucose metabolism and cytoskeleton alteration. Upon stimulation with high glucose, the expression of prominin-1 was up-regulated with concomitant down-regulation of its phosphorylation. Prominin-1 mutated at its phosphorylation site inflicted a global change in the expression of several genes associated with glucose metabolism in L6 myotube cells. Further, the over-expression of prominin-1 promoted glucose uptake in these cells. Prominin-1 undergoes sustained repression of its expression during confluence and differentiation of L6 myotube cells. The expression of prominin-1 in the MDCK cell modulated cell morphology and promoted cellular migration. These data imply that prominin-1 is involved in glucose metabolism and may regulate cellular glucose through its effect on cytoskeleton.

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