Interactions of DNA replication factors in vivo as detected by introduction of suppressor alleles of dnaA into other temperature-sensitive dna mutants

Abstract

Suppressor mutations located within dnaA can suppress the temperature sensitivity of a dnaZ polymerization mutant, indicating in vivo interaction of the products of these genes. The suppressor allele of dnaA [designated dnaA(SUZ, Cs)] could not be introduced, even at the permissive temperature, by transduction into temperature-sensitive (Ts) dnaC or dnaG recipients; it was transduced into dnaB(Ts) and dnaE(Ts) strains but at very low frequency. Recipient cells which were dnaA+ dnaE(Ts) were killed by the incoming dnaA(SUZ, Cs) allele, and it is presumed that combinations of dnaA(SUZ, Cs) with dnaB(Ts), dnaC(Ts), or dnaG(Ts) are lethal also. In one specific case, the lethality required the presence of three alleles: the incoming dnaA suppressor mutation, the resident dnaA+ gene, and the dnaB(Ts) gene. This was shown by the fact that dnaB(Ts) could readily be introduced into a dnaA(SUZ, Cs) dnaB+ recipient. That is, in the absence of dnaA+, the dnaA suppressor and dnaB(Ts) double mutant was stable. One model to explain these results proposes that the dnaA protein functions not only in initiation but also in the replication complex which contains multiple copies of dnaA and other replication factors.

Suppression of a DnaX temperature‐sensitive polymerization defect by mutation in the initiation gene, dnaA, requires functional oriC

“…(iv) Combination of the dnaA(Cs,Sx)73 mutation, A213D, with some dna gene Ts alleles was lethal. The Cs,Sx mutation could not be transduced into, or was transduced with very low efficiency into, dnaB, C, E, and G(Ts) mutants (Blinkova and Walker, 1983). (v) The protein(s) being suppressed, τ (and/or γ), as the replisome organizer (Yuzhakov et al ., 1996), also has multiple interactions within the replication complex, including possible association with the daughter chromosome partitioning machinery (Levine and Marians, 1998).…”

Physiological properties of cold-sensitive suppressor mutations of a temperature-sensitive dnaZ mutant of Escherichia coli

“…About one-third map within the dnaA initiation gene (42) and confer phenotypes expected of initiation-defective mutants. Properties of the suppressor-containing strains and additional evidence that dnaA protein interacts with other replication factors are reported here and in the accompanying paper (4).…”

“…Another line of evidence that dnaA and dnaZ proteins interact in vivo is presented in the accompanying paper (4). The dnaA(SUZ, Cs) suppressor mutation could not be transferred by transduction into dnaC(Ts) or dnaG(Ts) recipients.…”

Map location of the pcbA mutation and physiology of the mutant

“…This culture was then assayed for plaques by soft-agar overlay at 32 and 43°C. Ratios were determined for Table 2. Resulting titers were frequently less than on the optimal permissive host.…”