MUTATIONAL ANALYSIS OF THE VESICULAR STOMATITIS-VIRUS GLYCOPROTEIN-G FOR MEMBRANE-FUSION DOMAINS

The spike glycoprotein G of vesicular stomatitis virus (VSV) induces m embrane fusion at low pH. We used linker insertion mutagenesis to char acterize the domain(s) of G glycoprotein involved in low-pH-induced me mbrane fusion. Two or three amino acids were inserted in frame into va rious positions in the extracellular domain of G, and 14 mutants were isolated. All of the mutants expressed fully glycosylated proteins in COS cells. However, only seven mutant G glycoproteins were transported to the cell surface. Two of these mutants, D1 and A6, showed wild-typ e fusogenic properties. The mutant A2 had a temperature-sensitive defe ct in the transport of the mutant G glycoprotein to the cell surface. The other four mutants, H2, H5, H10, and A4, although present in cell surface, failed to induce cell fusion when cells expressing these muta nt glycoproteins were exposed to acidic pH. These four mutant G protei ns could form trimers, indicating that the defect in fusion was not du e to defective oligomerization. One of these mutations, H2, is within a region of conserved, uncharged amino acids that has been proposed as a possible fusogenic sequence. The mutation in H5 was about 70 amino acids downstream of the mutation in H2, while mutations in H10 and A4 were about 300 amino acids downstream of the mutation in H2. Conserved sequences were also noted in the H10 and A4 segment. The results sugg est that in the case of VSV G glycoprotein, the fusogenic activity may involve several spatially separated regions in the extracellular doma in of the protein.