Effect of recF, recJ, recN, recO and ruv mutations on ultraviolet survival and genetic recombination in a recD strain of Escherichia coli K12

Lloyd, Robert G.; Porton, Martin C.; Buckman, Carol

doi:10.1007/bf00334702

Cited by 104 publications

(76 citation statements)

References 38 publications

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“…The minimal medium was 56/2 salts supplemented as described by Low (19). 56/2 agar containing 40 ,ug of 5-bromo-4-chloro-3-indolyl phosphate (Xp) per ml was Hfr and F' matings and transductions with phage Plvir have been described elsewhere, as have methods for measuring recombination in Hfr crosses and sensitivity to UV light (16,18,21). Preparation and titration of stocks of phage A were according to the methods and media described by Silhavy et al (26).…”

Section: Methodsmentioning

confidence: 99%

Identification of sbcD mutations as cosuppressors of recBC that allow propagation of DNA palindromes in Escherichia coli K-12

1992

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The function of an open reading frame (orf-45) located upstream of the sbcC gene of Escherichia coli was investigated. Mutations that inactivate sbcC improve the ability to propagate Ared gam phage that carry a palindromic sequence in their DNA. They also act with sbcB mutations as cosuppressors of the defects in recombination, DNA repair, and cell viability associated with recBC mutations. A 1,282-bp cassette encoding resistance to kanamycin was used to disrupt orf-45. The mutation, which has a polar effect on the expression of sbcC, allowed stable propagation of palindromic k phage even when the sbcC gene product was provided in trans. Additional nonpolar mutations in orf-45 were isolated on the basis of their ability to improve the growth of recBC sbcB strains. These mutations also confer resistance to mitomycin C, allow efficient recombination in Hfr crosses, and facilitate stable propagation of palindromic phage. It is concluded that the products of orf-45 and sbcC are functionally related. The orf-45 gene is therefore renamed sbcD.The sbcC gene of Escherichia coli was defined originally by mutations that arise spontaneously in recBC sbcB strains (16). These mutations act with mutations in sbcB to suppress the defects in recombination and DNA repair associated with the recBC genotype and are selected, therefore, during normal growth because of the improved viability. Subsequently, sbcC single mutants were shown to be good hosts for a Xred gam phage carrying a DNA sequence with palindromic symmetry (Xpal) (5). Phage of this type grow poorly in sbcC+ cells. Leach and Stahl (13) had observed this feature of the sbcC phenotype in their earlier work with recBC sbcB sbcC strains but, because isogenic strains were not constructed and used for comparisons, had attributed it to the absence of exonuclease V (RecBCD enzyme) and exonuclease I (the product of sbcB). This seemed reasonable since palindromes might form hairpins or cruciforms which could be recognized and cleaved by nucleases that normally resolve Holliday intermediates in genetic recombination. Kulkarni and Stahl (11) MATERIALS AND METHODSBacterial strains and X phages. The E. coli K-12 strains and A phages used are described in Table 1. Phage stocks were prepared on strain JC7623, which allows stable propagation of the 571-bp palindrome carried by XMMS1632.Plasmids. Plasmid constructs carrying DNA inserts from the sbcC-orf-45 region of the chromosome are shown in Fig. 1. pSB43, pJP77, and pIN510 are described in detail elsewhere (3,23,28). pIN507 corresponds to pJP77 sbcC:: TnlOOO no. 5 described by Naom et al. (23). pFG103 was made by cloning a BamHI-HindIII fragment of approximately 14.5 kb from pJP77 into pACYC184 (6). A 1,282-bp cassette (kan) encoding resistance to kanamycin was cloned from pUC4K (29) into the unique PstI site of pFG103 to give pFG103K. pFG106 was constructed by removing the Kmr insertion from pFG103K by digestion with EcoRI and inserting it into the EcoRI site of the temperature-sensitive replicon pHSG415 (10). The sbcC+ construc...

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Section: Methodsmentioning

confidence: 99%

Identification of sbcD mutations as cosuppressors of recBC that allow propagation of DNA palindromes in Escherichia coli K-12

1992

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“…However, if the double mutant showed additional or synergistic effects then the pnp effect is mediated independently of that pathway (non-interacting genes). Such epistasis analysis has been extensively used to define interactions or non-interactions at the genetic level (Mahdi & Lloyd, 1989;Howard-Flanders et al, 1966;Cardenas et al, 2009;Lloyd et al, 1988;Li & Waters, 1998;Mandal et al, 1993).…”

Section: Discussionmentioning

confidence: 99%

Involvement of pnp in survival of UV radiation in Escherichia coli K-12

et al. 2012

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Polynucleotide phosphorylase (PNPase), a multifunctional protein, is a 39A59 exoribonuclease or exoDNase in the presence of inorganic phosphate (P i ), and extends a 39-OH of RNA or ssDNA in the presence of ADP or dADP. In Escherichia coli, PNPase is known to protect against H 2 O 2 -and mitomycin C-induced damage. Recent reports show that Bacillus subtilis PNPase is required for repair of H 2 O 2 -induced double-strand breaks. Here we show that absence of PNPase makes E. coli cells sensitive to UV, indicating that PNPase has a role in survival of UV radiation damage. Analyses of various DNA repair pathways show that in the absence of nucleotide excision repair, survival of UV radiation depends critically on PNPase function. Consequently, uvrA pnp, uvrB pnp and uvrC pnp strains show hypersensitivity to UV radiation. Whereas the pnp mutation is nonepistatic to recJ, recQ and recG mutations with respect to the UV-sensitivity phenotype, it is epistatic to uvrD, recB and ruvA mutations, implicating it in the recombinational repair process.

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“…Hfr donors were mated with F-recipients in high-salt LB broth by using procedures described in detail elsewhere (21,23). Matings were for 60 min at 37°C, with a donor-to-recipient ratio of 1:10. Colonies of recombinants were selected on 56/2 agar supplemented as described by Low (26) and containing 100 jig of streptomycin per ml to counterselect donor cells.…”

Section: Methodsmentioning

confidence: 99%

Linkage distortion following conjugational transfer of sbcC+ to recBC sbcBC strains of Escherichia coli

Lloyd

1991

J Bacteriol

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Conjugational recombination in Escherichia coli depends normally on RecBCD enzyme, a multifunctional nuclease and D)NA helicase produced by the recB, recC, and recD genes. However, recombination can proceed efficiently without RecBCD in recB or recC strains carrying additional mutations in both the sbcB and sbcC genes. Recombination in these strains, sometimes referred to as the RecF pathway, requires gene products that are not essential in the RecBCD-dependent process predominating in the wild type. It has also been reported to produce a different spectrum of recombinant genotypes in crosses with Hfr donors. However, the sbcC+ gene was unknowingly transferred to the recipient strain in some of these crosses, and this may have affected the outcome. This possibility was examined by conducting parallel crosses with Hfr donors that were either wild type or mutant for sbcC. Transfer of sbcC+ from an Hfr donor is shown to alter the frequency of recombinant genotypes recovered. There is a severe reduction in progeny that inherit donor markers linked to the sbcC+ allele and an increase in the incidence of multiple exchanges. Colonies of mixed genotype for one or more of the unselected proximal markers are also much more prevalent. Since the yield of recombinants is lower than normal, these changes are attributed to the reduced viability of recombinants that inherit sbcC+ from the Hfr donor. When the Hfr donor used is also mutant for sbcC, the yield of recombinants is greater and the frequencies of the different genotypes recovered are similar to those obtained in crosses with a rec+ sbc+ recipient, in which transfer of sbcC' has no apparent effect. Earlier studies are re-examined in light of these findings. It is concluded that, while recombination in recBC sbcBC strains involves different enzymes, the underlying molecular mechanism is essentially the same as that in the wild type.

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Effect of recF, recJ, recN, recO and ruv mutations on ultraviolet survival and genetic recombination in a recD strain of Escherichia coli K12

Cited by 104 publications

References 38 publications

Identification of sbcD mutations as cosuppressors of recBC that allow propagation of DNA palindromes in Escherichia coli K-12

Identification of sbcD mutations as cosuppressors of recBC that allow propagation of DNA palindromes in Escherichia coli K-12

Involvement of pnp in survival of UV radiation in Escherichia coli K-12

Linkage distortion following conjugational transfer of sbcC+ to recBC sbcBC strains of Escherichia coli

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