Transepithelial transport of a viral membrane glycoprotein implanted into the apical plasma membrane of Madin-Darby canine kidney cells. II. Immunological quantitation.
The envelope of vesicular stomatitis virus was fused with the apical plasma membrane of Madin-Darby canine kidney cells by low pH treatment. The fate of the implanted G protein was then followed using a protein A-binding assay, which was designed to quantitate the amount of G protein in the apical and the basolateral membranes. The implanted G protein was rapidly internalized at 31 degrees C, whereas at 10 degrees C no uptake was observed. Already after 15 min at 31 degrees C, a fraction of the G protein could be detected at the basolateral membrane. After 60 min 25-48% of the G protein was basolateral as measured by the protein A-binding assay. At the same time, 25-33% of the implanted G protein was detected at the apical membrane. Internalization of G protein was not affected by 20 mM ammonium chloride or by 10 microM monensin. However, the endocytosed G protein accumulated in intracellular vacuoles and redistribution back to the plasma membrane was inhibited. We conclude that the implanted G protein was rapidly internalized from the apical surface of Madin-Darby canine kidney cells and a major fraction was routed to the basolateral domain.
D000626:Aminopeptidases / Q000378:metabolism; D000818:Animals; D002451:Cell Compartmentation; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D004285:Dogs; D004533:Egtazic Acid / Q000494:pharmacology; D004705:Endocytosis; D004848:Epithelium / Q000378:metabolism; D006023:Glycoproteins / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007668:Kidney / Q000378:metabolism*; D008562:Membrane Glycoproteins; D008565:Membrane Proteins / Q000378:metabolism*; D013205:Staphylococcal Protein A; D014759:Viral Envelope Proteins; D014764:Viral Proteins / Q000276:immunology / Q000378:metabolism*
C028391:G protein, vesicular stomatitis virus; D006023:Glycoproteins; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D013205:Staphylococcal Protein A; D014759:Viral Envelope Proteins; D014764:Viral Proteins; D004533:Egtazic Acid; D000626:Aminopeptidases