Mutations ts2158 and ts1882, which confer temperature sensitivity of septum formation, map near leu in the region of min 2.0 to 2.1 on the Escherichia coli chromosome. These mutants stop division abruptly and grow as filaments at 42 C; when returned to 28 C, division resumes after about 30 min to produce short cells. The product of the gene defined by these mutations probably is required during all stages of septum formation rather than specifically for initiation of septation. Filaments that formed at 42 C contained incomplete constrictions (septa). When actively dividing filaments (i.e., those incubated at 28 C until division resumed) were shifted to 42 C a second time, division again stopped abruptly and incomplete constrictions persisted during the incubation at 42 C. Filaments that were subjected to 28 C incubation for a brief time (10, 20, or 30 min) before being shifted again to 42 C did not resume division as would be expected of a strain defective in initiating septation. Mutations ts1882 and ts2158 are recessive to the ts+ allele, which is consistent with the interpretation that these mutations cause the loss of a function. They did not complement each other and presumably represent one cistron. Mutants carrying ts1882 and ts2158 mutations were compared with a mutant defective in the ftsA allele, also known to map near leu.

Three mutants of Escherichia coli K-12 which form filaments during 42 C incubation have been characterized. The mutant strains AX621, AX629, and AX655 continued to grow and to synthesize deoxyribonucleic acid at 42 C for 150 to 180 min, after which time growth ceased. When cultures of the mutants were transferred from 42 to 28 C, septation of the filaments began after a 25to 30-min period and continued at a greater than normal rate until no filaments remained. Addition of chloramphenicol at the time of transfer from 42 to 28 C prevented cell division in strain AX655 and caused lysis of strains AX621 a;;d AX629. The temperature sensitivity mutation in each strain mapped near leu. For strain AX621, the mutation was specifically located between leu and nadC by P1 transduction. Properties of these strains are compared with those of other cell division mutants.

The dnaH locus is the fourth gene to be identified as required for deoxyribonucleic acid polymerization in Escherichia coli. A temperature-sensitive mutant defective in this gene exhibited an abrupt decrease in rate of deoxyribonucleic acid synthesis when shifted to 42 C. The locus mapped in the proC-purE region of the chromosome by conjugation and was co-transducible with purE. dnaH+ is carried on the F',8 episome and is dominant over the dnaHmutation.Mutations which cause temperature sensitivity of deoxyribonucleic acid (DNA) replication in Bacillus subtilis and Escherichia coli (11) have been described. These mutations have been mapped at nine loci in B. subtilis (19) and at six loci in E. coli (36,37). In E. coli, the dnaA and dnaC gene products are required for replication initiation, dnaB, dnaE; and dnaG gene products are involved in polymerization (4,7,15,16,20,21), and dnaF (nrdA) is involved in precursor biosynthesis (8).We describe here an additional mutant of E. coli which is temperature sensitive for DNA synthesis and cell division. Upon a temperature shift to 42 C, rate of DNA synthesis and of cell division decreased immediately, but ribonucleic acid (RNA) synthesis and growth continued for two or three doublings. Viability decreased exponentially after a lag of about 20 min. The temperature sensitivity mutation was co-transducible with purE, and the locus is designated dnaH. Partial diploid strains which carry dnaH+ on an episome and dnaH on the chromosome were able to synthesize DNA, divide, and form colonies at 42 C; therefore, the wild-type dnaH allele is dominant. MATERIALS AND METHODSStrains. The principal bacterial strains used are listed in Table 1. Origins of the Hfr strains are shown in Fig. 1. Bacteriophages Plvir and f2 were obtained from J. L. Rosner and R. Johnston, respectively. Growth conditions. The yeast extract-tryptone medium of Howard-Flanders et al. (18) with 0.5% NaCl was used unless stated otherwise. Low-phosphate medium was the defined medium base (18) containing 0.1 M tris(hydroxymethyl)aminomethane (pH 7.2) in place of phosphate buffer and supple-mented with the following: glucose, 10 mg/ml; thiamine-hydrochloride, 5 ug/ml; Casamino Acids (Difco), 2.5 mg/ml; and phosphate, 0.2 gsmol/ml. Incubation was at 28 or 42 C with shaking. Temperature shifts were accomplished by transfer of cultures to previously equilibrated flasks.Phages f2 and Plvir were grown by the procedures of Loeb and Zinder (23) and Rosner (32), respectively.Isolation of the temperature-sensitive mutant AX642. Strain AX642, which carries the temperaturesensitive (ts) mutation ts20-16, was isolated during a process of isolating conditional cell division mutants. After mutagenesis of E. coli K-12 AB1157 (2) and screening of survivors for inability to form colonies at 42 C, temperature-sensitive clones were screened further for filament formation at 42 C. Mutant AX642 was discovered because it formed filaments which were three to four times the normal cell length during incubation at 42 C.Cell number, mass, and ...

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