B. GP44, gp50, and gp51 regulate the shutoff of host biosyntheses indirectly, by regulating expression of the genes of the host-takeover module.GPS 44/50/51 target RNA polymerase (RNAP). SPO1 gene 44 (then known as e3) was cloned in a B. subtilis:E. coli shuttle vector in which the cloned gene is expressed from an inducible promoter. When expressed in uninfected cells of either B. subtilis or E. coli, it caused the shutoff of bacterial RNA synthesis and cell death. GP44 and gp51 show extensive homology, including acidic/hydrophobic domains similar to the domain in E. coli σ54 that is required for binding to RNAP (8). Because of the very small size of gp50 (23 amino acids), the overlap between genes 50 and 51, and the synergistic effect of mutations in genes 50 and 51, we suppose that gp50 acts jointly with gp51 (6,4). Thus, it seems likely that gp50/51 also targets the host RNAP.
Roles of gp44, gp50, and gp51 in host-shutoff. Shutoff of host RNA synthesis was accelerated by either the 44- single mutant or the 50-51- double mutant, and more so by the 44-50-51- triple mutant. Shutoff of host DNA synthesis was accelerated by the mutants early in infection but delayed by the 44-50-51- triple mutant at late times. These observations suggested that the role of gp44/50/51 in host-shutoff is regulatory, rather than directly causative. In fact, gp44/50/51 does regulate the expression of SPO1 genes involved in host-shutoff. Either gp44 alone or gp50/51 alone represses immediate-early genes and stimulates expression of middle and delayed-early genes. When all three proteins are present together the effect on immediate-early genes is increased, the effect on delayed-early and middle genes is decreased, and the effect on some delayed-early genes is reversed. These effects on regulation of gene expression provide a plausible explanation for the effects on host shutoff. The accelerated shutoff of host RNA synthesis by the triple mutant could be explained if that shutoff were caused by the product(s) of immediate-early gene(s) that are overexpressed in infection by the triple mutant. The deficient shutoff of host DNA synthesis by the triple mutant at middle times could be explained if that shutoff required middle gene product(s) whose expression is deficient in the triple mutant. (4). Thus, our present belief is that the binding of gp44/50/51 to RNAP is primarily for the purpose of regulating phage gene expression, and that the shutoff of host RNA synthesis, that occurs when gp44 or gp50/51 is expressed in uninfected cells, is an incidental consequence of their binding to RNAP.