Interactions of wild-type and mutant M protein of vesicular stomatitis virus with viral nucleocapsid and envelope in intact virions. Evidence from [125I]iodonaphthyl azide labeling and specific cross-linking.
Four different temperature-sensitive M protein mutants (tsM) of vesicular stomatitis virus (VSV) were characterized with regard to the association of the mutated M protein either with nucleocapsids or with membranes in the intact virions. Virions were labeled with the photoreactive hydrophobic probe [125I]iodonaphthyl azide (INA) to assess interactions between viral proteins and the lipid envelope. In wild type (wt) virions, the three major structural proteins--G, M, and N--were labeled in the ratio ca. 1.0:0.4:0.2. INA labeled only the membrane-associated peptide of G protein, both in the intact virion and in reconstituted G protein--viral lipid vesicles, demonstrating the specificity of INA for lipid bilayer regions. Labeling of tsM virions with INA resulted in a 2--3-fold greater incorporation into M protein than was found for wt virions, suggesting increased M--membrane associations in the mutant virions. Temperature-stable revertants from tsM possessed wt labeling characteristics. Interaction of the M protein with nucleocapsids was assessed from the abundance of disulfide-linked M--N complexes found after disruption of the virions by sodium dodecyl sulfate solution under nonreducing conditions. The abundance of such complexes was 30--80% less from tsM virions than from wt virions, suggesting decreased M--nucleocapsid interactions in tsM virions. Temperature-stable revertants from tsM resembled wt in the abundance of M--N complex formed. We conclude that the mutations alter M protein in such a way as simultaneously to increase its association with membrane and to decrease its affinity for nucleocapsids in the intact virion.
D000818:Animals; D001386:Azides / Q000494:pharmacology*; D002460:Cell Line; D006224:Cricetinae; D004279:DNA, Viral / Q000378:metabolism*; D007457:Iodine Radioisotopes; D007668:Kidney; D007700:Kinetics; D009154:Mutation; D009281:Naphthalenes / Q000494:pharmacology*; D014721:Vesicular stomatitis Indiana virus / Q000378:metabolism*; D014763:Viral Matrix Proteins; D014764:Viral Proteins / Q000378:metabolism*; D014771:Virion / Q000378:metabolism*
D001386:Azides; D004279:DNA, Viral; D007457:Iodine Radioisotopes; D009281:Naphthalenes; D014763:Viral Matrix Proteins; D014764:Viral Proteins; C015298:iodonaphthylazide