Incorporation of a charged amino acid into the membrane-spanning domain blocks cell surface transport but not membrane anchoring of a viral glycoprotein.
The membrane-spanning domain of the vesicular stomatitis virus glycoprotein (G protein) consists of a continuous stretch of 20 uncharged and mostly hydrophobic amino acids. We examined the effects of two mutations which change the amino acid sequence in this domain. These mutations were generated by oligonucleotide-directed mutagenesis of a cDNA clone encoding the G protein, and the altered G proteins were then expressed in animal cells. Replacement of an isoleucine residue in the center of this domain with a strongly polar but uncharged amino acid (glutamine) had no effect on membrane anchoring or transport of the protein to the cell surface. Replacement of this same isoleucine residue with a charged amino acid (arginine) generated a G protein that still spanned intracellular membranes but was not transported efficiently to the cell surface. The protein accumulated in the Golgi region in about 50% of the cells, and about 20% of the cells had detectable protein levels in a punctate pattern on the cell surface. In the remaining cells the protein accumulated in a vesicular pattern throughout the cytoplasm. Models which might explain the abnormal behavior of this protein are discussed.
Adams
G A
GA
Rose
J K
JK
D000595:Amino Acid Sequence; D000818:Animals; D001692:Biological Transport; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D006023:Glycoproteins / Q000235:genetics / Q000378:metabolism*; D007532:Isoleucine / Q000502:physiology; D008562:Membrane Glycoproteins; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D011487:Protein Conformation; D011499:Protein Processing, Post-Translational; D014721:Vesicular stomatitis Indiana virus / Q000235:genetics*; D014759:Viral Envelope Proteins; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*
C028391:G protein, vesicular stomatitis virus; D006023:Glycoproteins; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D014759:Viral Envelope Proteins; D014764:Viral Proteins; D007532:Isoleucine