FIG. 1.

Myristoylation of arenavirus Z protein is required for VLP formation. (A) Alignment of the N termini of LCMV and LFV Z protein sequences. The arrow indicates the conserved G2 residue. Basic amino acid residues are underlined. (B) Expression levels of wt and mutant G2A Z proteins. 293T cells were transfected with pCAGGS plasmids coding for the indicated protein. Cell lysates were prepared 48 h after transfection and were analyzed by Western blot using an anti-HA antibody. (C) Mutation G2A eliminates VLP formation. 293T cells were transfected with the indicated combination of plasmids. Forty-eight hours later, SP were saved and cell lysates were prepared for CAT assay. An aliquot of each SP was used to infect a fresh monolayer of BHK-21 cells, which were subsequently infected with LCMV (MOI = 3). Seventy-two hours postinfection cell lysates were prepared for CAT assay. Aliquots from lysates of 293T (transfection) and BHK-21 (passage) cells were assayed for CAT activity. NAc, nonacetylated chloramphenicol; MAc, monoacetylated chloramphenicol. (D) wt, but not G2A, Z protein is myristoylated. COS-1 cells were transfected with plasmid expressing wt or G2A mutant Z proteins, and 44 h later they were metabolically labeled with either [³⁵S]methionine-cysteine (³⁵S) or [9,10 (n)-³H]myristic acid (³H) for 4 h. Z proteins were immunoprecipitated from cell extracts by using an anti-HA antibody. Immunoprecipitates were resolved by SDS-PAGE. The gel was then subjected to fluorography, dried, and exposed.