| The plasma membrane of polarized epithelial cells is divided into apical and basolateral domains, each composed of a unique set of proteins and lipids. In order to establish and maintain polarity in these cells, apical and basolateral membrane proteins are sorted along the biosynthetic pathway, as well as along the endocytic pathway during recycling to the plasma membrane. Sorting to the basolateral surface is mediated by distinct targeting signals found within the cytoplasmic tail of transmembrane proteins. The aim of this thesis is to define precisely the pathways for the transport of select membrane proteins to the basolateral surface of polarized epithelial cells.; We have shown that the proteoglycan, syndecan-1, is targeted to the basolateral membrane by a type II PDZ binding motif. These PDZ-mediated interactions contribute, in a novel fashion, to the polarized sorting of newly synthesized syndecan-1 prior to arrival at the cell surface. Using RNA interference, we attempted to identify the PDZ protein(s) that sorts syndecan-1 to the basolateral membrane.; The polarized transport of many membrane proteins to the basolateral surface is thought to be dependent on the epithelial specific clathrin adaptor complex, AP-1B. The mechanism by which AP-1B sorts basolateral cargo, however, remains unclear. To precisely define the targeting signals interpreted by AP-1B, we used RNA interference to specifically deplete the mu1B subunit. We find that basolateral signals dependent on a tyrosine also used for clathrin-mediated endocytosis are decoded by AP-1B. Additionally, since AP-4 has also been implicated in polarized transport, we used RNA interference to examine the contribution of AP-4 individually, and in combination with AP-1B, to basolateral targeting. Our results show that basolateral targeting cannot be completely explained by the activities of clathrin adaptor complexes, either singly or together. Although some sorting events are clearly dependent on this class of adaptor, it is now clear that other such sorting elements remain to be discovered. |
