The Role of D-Cbl in Epithelial Cellpolarity

Cell polarity is a basic feature of eukaryotic cells from budding yeast to mammalian epithelia. The evolutionarily conserved mechanisms have been applied for the generation and maintenance of cell polarity. Loss of cell polarity and tissue architecture are characteristics of malignant cancers derived from epithelial tissues. The germ cells in Drosophila egg chamber are surrounded by a follicular epithelial sheet, which is characterized by columnar shape, strong cell-cell adhesion, and pronounced apicobasal polarity. Therefore, the Drosophila follicle cells provide an ideal in vivo system to study cell polarity. Our lab has being focused on functional studies of D-cbl in Egfr signaling for years. Recently, we have observed an intriguing phenotype on mutant follicle cell clones for D-cbl, which displayed a loss of epithelial cell polarity. Mutant cells were round, showing a multilayered effect, and sometime invaded into the germ cell area. We also noticed that D-CblS contains a PDZ binding motif, which may mediate an interaction with PDZ proteins. PDZ proteins are known for retaining and regulating membrane transporters in polarized epithelial cell membranes. We suspect that D-CblS, as an adaptor, may form complex with PDZ proteins, and they together determine epithelial cell polarity. We went on to test this possibility by disrupting this motif. Interesting, if this motif is mutated in D-CblS, the lateral membrane localization of D-CblS was lost. Ectopic expression of this mutant form of D-CblS resulted in lethal effect, and the loss of follicle cell polarity indicated by the mis-localization of E-cadherin, and invasive behavior. Furthermore, we found that the lateral membrane localization of D-CblS was disrupted in scrib heterozygote mutant, but not in bazooka or crumbs mutants. Here we proposed to dissect the role of D-CblS in epithelial cell polarity. First of all, we ask what is the molecular mechanism that underpins the cell polarity function of D-cbl? Examining the effect of lack of D-Cbl on the distribution of junctional proteins, such as the apical Par complex and components of adherens junction, Armadillo, will provide molecular mechanisms for the loss of cell polarity effects. We then will use yeast two hybrid system to identify the D-CblS interacting PDZ proteins, which might be Scrib or proteins in Scrib complex, like Dgl or Lgl. The effect of D-CblS on the Scrib complex will be analyzed by looking at the interaction between Scrib complex and the exocyst, which is known for mediating exocytosis at specific sites of the plasma membrane that is critical for basolateral membrane proteins. We will also try to elucidate what is the pathway upstream of D-cbl that decides where and when it functions? We will test whether Egfr signaling is required for cell polarity maintenance by D-CblS.

Project ID:PA9902-0325
External Project ID:NSC97-2311-B182-001-MY3