The role of gene V protein in f1 single-strand synthesis

Abstract

We demonstrate that, in cells infected with phage f1, only the gene V protein, a DNA-binding protein, is necessary to effect the switch from double- to single-strand synthesis; we specifically show that no host proteins are required for single-strand synthesis beyond those that are already required for double-strand synthesis. f1 temperature-sensitive ( ts) gene II-infected cells were allowed to accumulate excess gene V protein in the absence of DNA replication through incubation at restrictive temperature. The infected cells were then shifted to permissive temperature in the presence of chloramphenicol; single-strand synthesis ensued. When, however, the identical experiment was performed with wild-type f1-infected cells, double, rather than single strands were synthesized. Since the principal difference between the two experiments lay in the accumulation of excess gene V protein in the tsII infection, single-strand synthesis must have halted in the wild-type infection because no free gene V protein was available and not because a chloramphenicol-sensitive host protein was being depleted. Chloramphenicol, by preventing protein synthesis, apparently blocks the two sources of gene V protein normally used for single-strand synthesis. No new gene V protein can be synthesized, and all previously synthesized gene V protein remains stably complexed with preexisting single strands rather than being recycled during morphogenesis of the single strands. In the absence of both new and recycled gene V protein, double-strand synthesis resumes. If a failure to recycle gene V protein were indeed the lesion induced by chloramphenicol treatment, then inactivation of specific f1 gene products required for morphogenesis, through use of amber and temperature-sensitive mutant phage, might also lead to replacement of single-strand synthesis by double-strand synthesis. Such cessation of single-strand synthesis was indeed found after infections by mutants in genes I, IV, VII, and VIII but not after infections by mutants in genes III and VI. The gene III and VI proteins therefore act at a later step in morphogenesis than the gene I, IV, VII, and VIII proteins; they appear to function after the single strands and gene V protein have separated. Data obtained from temperature shift experiments indicate that the supply of gene V protein in an infected cell is carefully regulated and that there is never much free gene V protein available.