Homophilic adhesion of the myelin P0 protein requires glycosylation of both molecules in the homophilic pair.
The myelin P0 protein is glycosylated at a single site, asparagine 93, within its only immunoglobulin (Ig)-like domain. We have previously shown that P0 behaves like a homophilic adhesion molecule (Filbin, M. T., F. S. Walsh, B. D. Trapp, J. A. Pizzey, and G. I. Tennekoon. 1990. Nature (Lond.). 344:871-872). To determine if the sugar residues of this molecule contribute to its adhesiveness, the glycosylation site was eliminated by replacing asparagine 93 with an alanine, through site-directed mutagenesis of the P0 cDNA. The mutated P0 cDNA was transfected into CHO cells and surface expression of the mutated P0 was assessed by immunofluorescence, limited trypsinization and an ELISA. A cell line was chosen which expressed approximately equivalent amounts of the unglcosylated P0 (UNGP0) at the cell surface as did a cell line expressing the fully glycosylated P0 (GPo); the adhesive properties of these two cell lines were compared. It was found that when a single cell suspension of the UNGPo cells were incubated, by 60 min, unlike the GP0 cells, they had not formed large aggregates; they were indistinguishable from the control transfected cells. This suggests that the UNGP0 protein does not behave like an adhesion molecule. To establish if only one molecule in the P0:P0 homophilic pair must be glycosylated for adhesion to occur, the ability of UNGP0 cells to adhere to GP0 cells was assessed both qualitatively and quantitatively. The results of both types of assay imply that, indeed, both P0 molecules in the homophilic pair must be glycosylated for adhesion to take place.
Filbin
M T
MT
Department of Biological Sciences, Hunter College of the City University of New York, New York 10021.
Tennekoon
G I
GI
D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D002448:Cell Adhesion; D015816:Cell Adhesion Molecules, Neuronal / Q000235:genetics / Q000378:metabolism*; D006224:Cricetinae; D006031:Glycosylation; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D018993:Myelin P0 Protein; D009185:Myelin Proteins / Q000235:genetics / Q000378:metabolism*; D009838:Oligodeoxyribonucleotides
D015816:Cell Adhesion Molecules, Neuronal; D018993:Myelin P0 Protein; D009185:Myelin Proteins; D009838:Oligodeoxyribonucleotides