Biosynthesis of a phosphatidylinositol-glycan-linked membrane protein: signals for posttranslational processing of the Ly-6E antigen.
The Ly-6E/A protein is a murine cell surface protein expressed at high levels on activated peripheral T cells. The only linkage known to be responsible for its association with the plasma membrane is a phosphatidylinositol-glycan (PI-G) moiety. To examine the biosynthesis of this structure, we constructed a series of mutants of Ly-6E that were expressed in COS cells by using transient-transfection procedures. When 12 or 20 carboxy-terminal residues were deleted from the primary translation product, the PI-G modification was completely abolished and the mutant proteins became secreted. Addition of the PI-G tail was partially inhibited when the charged 12-amino-acid peptide found as a cytoplasmic tail on the transmembrane form of LFA-3 was added to the COOH terminus of the Ly-6E protein. Proteolytic cleavage occurred on this mutant protein, but the PI-G moiety was added to only 50% of the molecules. Changing an Asn residue to a Lys at the hypothetical cleavage site resulted in a PI-G-linked protein having a detectable alteration in electrophoretic mobility. This finding raises the possibility that proteolytic cleavage at other amino acid sites may occur and that PI-G attachment can occur at this new site. A model identifying two regions that may act as necessary signals for the biosynthesis of the PI-G tail is presented.
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Department of Pathology, Yale University Medical School, New Haven, Connecticut 06510.
Bothwell
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D000595:Amino Acid Sequence; D000950:Antigens, Ly / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D002478:Cells, Cultured; D004252:DNA Mutational Analysis; D005822:Genetic Vectors; D017261:Glycosylphosphatidylinositols; D008562:Membrane Glycoproteins / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D010716:Phosphatidylinositols / Q000378:metabolism*; D011134:Polysaccharides / Q000378:metabolism*; D011487:Protein Conformation; D011499:Protein Processing, Post-Translational
D000950:Antigens, Ly; D017261:Glycosylphosphatidylinositols; D008562:Membrane Glycoproteins; D010716:Phosphatidylinositols; D011134:Polysaccharides