Membrane topology of Bves/Pop1A, a cell adhesion molecule that displays dynamic changes in cellular distribution during development.

Knight RF, Bader DM, Backstrom JR
J Biol Chem. 2003 278 (35): 32872-9

PMID: 12815060 · DOI:10.1074/jbc.M301961200

We investigated the membrane topology of Bves/Pop1A as a foundation to dissect the molecular basis and function of Bves/Pop1A trafficking during development. Bves contains two asparagine-linked glycosylation sites within the amino terminus and three putative membrane domains. Therefore, glycosylation assays were performed to determine if the amino terminus of Bves is delivered into the endoplasmic reticulum lumen and glycosylated. We establish that Bves from chick heart and transfected cells is glycosylated, implying that the amino terminus of cell surface molecules is extracellular. Three biochemically distinct approaches were utilized to determine the orientation of the carboxyl terminus of Bves. First, glycosylation of Bves at exogenous sites within the carboxyl terminus was only observed in a construct that lacked the third membrane domain, which presumably reversed the orientation of the carboxyl terminus. Second, co-expression of full-length Bves with soluble, carboxyl-terminal Bves constructs that reside in different subcellular compartments revealed that Bves-Bves interactions occur in the cytoplasm. Third, the immunoreactivity of endogenous Bves at the cell surface of epicardial cells was dramatically enhanced with detergent. These results suggest that the membrane topology of cell surface Bves/Pop1A is composed of an extracellular amino terminus, three transmembrane domains, and a cytoplasmic carboxyl terminus. We therefore hypothesize that the carboxyl terminus regulates the cellular distribution of Bves/Pop1A during coronary vessel development.

MeSH Terms (32)

3T3 Cells Amino Acid Sequence Animals Asparagine Avian Proteins Biotinylation Cell Adhesion Cell Adhesion Molecules Cell Line Cell Membrane Chick Embryo COS Cells Cytoplasm Detergents Electrophoresis, Polyacrylamide Gel Endoplasmic Reticulum Gene Deletion Gene Expression Regulation, Developmental Glycosylation Immunohistochemistry Mice Models, Biological Models, Genetic Molecular Sequence Data Muscle Proteins Myocardium Precipitin Tests Protein Structure, Tertiary Rats Sequence Homology, Amino Acid Tissue Distribution Transfection

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