Alfalfa mosaic virus temperature-sensitive mutants. II. Early functions encoded by RNAs 1 and 2

Mutants Bts 03 and Mts 04 of alfalfa mosaic virus (AIMV) have temperature-sensitive mutations in genomic RNAs 1 and 2, respectively. These mutants are defective in the production of viral minus-strand RNA, coat protein, and infectious virus when assayed in cowpea protoplasts at the nonpermissive temperature (30 degrees). To determine the temperature-sensitive step in the replication cycle, mutant-infected protoplasts were shifted from an incubation temperature of 25 degrees (permissive temperature) to 30 degrees at different times during a 24-hr incubation period. For both mutants an initial incubation of infected protoplasts for 6 hr at 25 degrees was sufficient to permit a normal minus-strand RNA synthesis, translation of RNA 4 into coat protein, and assembly of infectious virus during the subsequent incubation at the nonpermissive temperature. Probably, AIMV RNAs 1 and 2 encoded proteins are produced early in infection and the mutant proteins are protected from inactivation at 30 degrees once they are incorporated in a functional structure.