| The cellular phosphoinositide signaling system, an intricate network of enzymes and phospholipid messengers, is a critical regulator of most, if not all, cellular processes. The cell generates and is able to utilize multiple phosphoinositides as unique messenger molecules in the transduction of extracellular or intracellular signaling. One of these messengers, phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2), is a potent regulator of a variety of cellular processes, including actin dynamics, cell adhesion, endocytic and exocytic events, and gene expression. However, Ptdlns(4,5)P2 must be generated in a site-specific manner in order to limit its cellular effects. By the spatial and temporal targeting of the phosphatidylinositol 4-phosphate, 5-kinases (PIPKs) via the actions of its functional protein associates, PtdIns(4,5)P2 is generated at discrete cellular locales where it is needed for cellular function. To facilitate this site-directed activity, the type I gamma PIPK (PIPKIgamma) is alternatively spliced on its extreme C-terminus. Here, I have identified and characterized two novel PIPKIgamma splice variants, PIPKIgamma_i4 and PIPKIgamma_i5, that are expressed in human cells and display distinct subcellular localization. As the targeting of PIPKs is a mechanism for signaling specificity, the function of PIPKIgamma_i5 in endosomal trafficking was explored. I identified that PIPKIgamma_i5 associates directly with sorting nexin 5 (SNX5), a regulator of intracellular trafficking pathways. SNX5 is a PtdIns(4,5)P2-binding protein, and accordingly, the association between PIPKIgamma_i5 and SNX5 was found to be regulated by PtdIns(4,5)P2 generation. This association was also regulated by tyrosine phosphorylation of PIPKIgamma_i5, suggesting that upstream signaling pathways can potentiate the PIPKIgamma_i5-SNX5 interaction. Importantly, the association between PIPKIgamma_i5 and SNX5 was found to regulate the lysosomal targeting of the cellular adhesion receptor E-cadherin. The combined data suggest a model where growth factor stimulation results in increased lysosomal targeting of E-cadherin via a PIPKIgamma_i5 -- mediated endosomal pathway, whereas SNX5 functions to antagonize PIPKIgamma_i5 promotion of E-cadherin degradation. Hence, the regulation of SNX5 activity by PIPKIgamma_i5 and PtdIns(4,5)P2 synthesis may serve as a crucial regulator of the epithelial--to-mesenchymal transition. |
