Identification of the pifC gene and its role in negative control of F factor pif gene expression

Abstract

The pif region of the F factor includes two genes, pifA and pifB, that lead to abortive T7 infection. We have identified a new gene in this region, pifC, by constructing an in vitro fusion of pif DNA at 41.6 kilobases on the F factor physical map to the lacZ gene. A PifC-LacZ fusion protein of 149,000 daltons has been identified by immunoprecipitation and polyacrylamide gel electrophoresis. This allows us to assign the N terminus of pifC to 42.5 kilobases on the F map. Using fusions of pifC, pifA, and pifB to lacZ, we have studied the regulation of pif gene expression and have shown that the product of pifC negatively controls its own expression and that of pifA and pifB.

F factor inhibition of conjugal transfer of broad-host-range plasmid RP4: requirement for the protein product of pif operon regulatory gene pifC

“…Phage T7 was originally from F. W. Studier. Solid and liquid growth media (ML) have been described previously (13,15).…”

“…The positions of the pGS103 mutations that resulted in PifC-phenotypes were determined by recombination with F'lac derivatives F'C521, B9, and 05, which contain overlapping deletions that join the pifC structural gene to lacZ. F'C521 carries a fusion between pifC, at 41.5 kilobases (kb), and the N terminus of lacZ and has been previously described in detail (13). 05 carries a pifC-lacZ fusion which retains only a small N-terminal portion of the pifC coding sequences (M. Malamy, unpublished data) fused to the distal portion of the a region of lacZ (8).…”

“…In contrast, RP4 has no effect on the efficiency of F factor transfer. By analyzing plasmids containing various segments of the F factor, Tanimoto and Iino (18) localized the sequences responsible for inhibition of RP4 transfer to the same region in which we mapped the pifC gene (13). In this report we examine whether inhibition of RP4 transfer is due to the pifC protein product (PifC), an RNA species transcribed in the pifC region, or a closely linked gene that requires the pif promoter for expression.…”

“…The F factor pifC gene is located in the immediate vicinity of oriVl (13), the primary origin of mini-F vegetative replication (5,6). The pifC gene product plays several roles: it is involved in the regulation of pif gene expression and the initiation of F factor DNA replication (13,14,19). We have shown previously that the pifC structural gene is contained within the pif operon, which includes two distal genes, pifA and pijB.…”

“…We have shown previously that the pifC structural gene is contained within the pif operon, which includes two distal genes, pifA and pijB. The protein products of the pifA and pifB genes inhibit development of bacteriophage T7 (3,16,17), and the product of the pifC gene functions as an autoregulatory control element which negatively regulates expression of the operon (13). The pifC product has been implicated by Tanimoto and lino (19) as a requirement for replication of an oriVI-dependent mini-F replicon.…”