Gene product 0.4 increases bacteriophage T7 competitiveness by inhibiting host cell division

Abstract

Bacteriophages take over host resources primarily via the activity of proteins expressed early in infection. One of these proteins, produced by the Escherichia coli phage T7, is gene product (Gp) 0.4. Here, we show that Gp0.4 is a direct inhibitor of the E. coli filamenting temperature-sensitive mutant Z division protein. A chemically synthesized Gp0.4 binds to purified filamenting temperature-sensitive mutant Z protein and directly inhibits its assembly in vitro. Consequently, expression of Gp0.4 in vivo is lethal to E. coli and results in bacteria that are morphologically elongated. We further show that this inhibition of cell division by Gp0.4 enhances the bacteriophage's competitive ability. This division inhibition is thus a fascinating example of a strategy in bacteriophages to maximize utilization of their hosts' cell resources.host takeover | bacterial division | bacteriophage biology T he abundance of bacteriophages and their importance to microbial evolution, and consequently to major ecological issues, provide an incentive to study their biology. Bacteriophage T7 and its host, Escherichia coli, provide a model for systematically studying host-virus interactions. The genetics of both the phage and its host have been studied extensively, and the putative functions or tentative physiological roles of over half of the 56 genes of phage T7 and the 4,453 genes of E. coli have been identified (1-3). T7 is a virulent phage which, upon infection of its host E. coli, produces over 100 progeny phage per host in less than 25 min. It is an obligatory lytic phage, meaning that a successful phage growth cycle always results in lysis of the host. The genome of T7 is a 39,937-bp double-stranded DNA molecule (4). Despite extensive knowledge of phage T7, the mechanism by which it takes over the host molecular machinery remains obscure. Specific functions have been attributed to over

Revealing bacterial targets of growth inhibitors encoded by bacteriophage T7

“…Twelve mutants resistant to this growth inhibitor were isolated, and their genome was deep-sequenced. We expected to identify a specific mutation that renders FtsZ refractory to Gp0.4 inhibition and hence enables survival of the bacteria encoding this mutation, as we previously reported (9). Indeed, following analysis of the sequencing results (described in the next section), we identified a unique mutation encoded only by resistant mutants expressing Gp0.4.…”

“…1). To test the feasibility of this approach, we expressed the growth inhibitor Gp0.4 of T7 bacteriophage, which inhibits the division protein FtsZ (9). Twelve mutants resistant to this growth inhibitor were isolated, and their genome was deep-sequenced.…”

“…Other genes were also mutated in these resistant mutants, but none were unique, as they were also mutated in mutants resistant to other growth inhibitors. The specific mutation was an insertion mutation of 6 nt in ftsZ, shown to confer resistance to Gp0.4 expression (9). The frequency of this mutation was 61.54%, suggesting it appears in the majority of the 12 resistant mutants ( Table 1).…”

“…Nevertheless, under other as-yet unidentified conditions, this gene's inhibition might be useful for phage growth. It is intriguing that the newly identified inhibitor of MreB, Gp0.6, is encoded near Gp0.4 (203 bp away), a recently identified inhibitor of FtsZ (9). MreB and FtsZ are key cytoskeletal proteins in E. coli.…”

“…Phages have coevolved with bacteria for over 3 billion years and have thus developed molecules to specifically and optimally inhibit or divert key metabolic functions. Examples of bacterial targets inhibited by phage-derived products include the δ subunit of the DNA polymerase III clamp loader, inhibited by gene product (Gp) 8 of the coliphage N4 (7); the Staphylococcus aureus putative helicase loader, DnaI, inhibited by ORF104 of bacteriophage 77 (5); a key enzyme of folate metabolism, FolD, inhibited by Gp55.1 of the coliphage T4 (8); and the essential cell-division protein, filamenting temperature-sensitive mutant Z (FtsZ), inhibited by Gp0.4 of the coliphage T7 (9). These examples suggest that there are other phage products that may inhibit other bacterial targets.…”