A SEC1-RELATED VESICLE-TRANSPORT PROTEIN THAT IS EXPRESSED PREDOMINANTLY IN EPITHELIAL-CELLS

Sec1-related proteins are involved in docking and fusion of transport vesicles in eukaryotic cells. Here we report the cloning and molecular characterization of a Sec1-related protein expressed in the MDCK epit helial cell line. This protein represents a canine counterpart of the murine Munc-18-2/Munc-18b/muSec1 protein, displays 93 % amino acid ide ntity with these proteins, has a similar tissue mRNA expression patter n, and associates in vitro with syntaxins 1A, 2, and 3. In situ hybrid ization analysis of embryonic mouse tissues revealed prominent express ion of the munc-18-2 mRNA in the epithelia of several tissues, Cell-fr actionation studies demonstrated that the majority of Munc-18-2 is mem brane associated, Most of the protein is washed off the membranes by s odium carbonate, pH 11.5, However, the protein is poorly solubilized b y detergent treatment. The Munc-18-2 protein was localized, by immunof luorescence microscopy, to the plasma membrane of MDCK cells, and is a pically distributed in the epithelial cells of mouse tissues, When ove rexpressed in COS-L cells, the protein appeared to be largely cytosoli c. However, upon expression with syntaxin 1A, it displayed a shift to the plasma membrane, where the two proteins colocalized. These results identified Munc-18-2 as a predominantly epithelial vesicle-transport protein with a polarized distribution and provided novel vivo evidence for the association of Sec1-related proteins with members uf the synt axin family.