Localization of the plasmid (pKM101) gene(s) involved in recA + lexA +-dependent mutagenesis

Shanabruch, William G.; Walker, Graham C.

doi:10.1007/bf00425456

Cited by 111 publications

(42 citation statements)

References 31 publications

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“…The phenotype of enhancing MMS mutagenesis but not increasing resistance to UV killing makes pGW16 different from the other pKM101 mutants that have been isolated to date (27,33,41), all of which have lost the abilities both to enhance MMS mutagenesis and to increase resistance to UV killing. However, a possible clue to the molecular basis of this unusual phenotype was provided by our observation that these effects caused by the presence of pGW16 in an E. coli cell were reminiscent of those caused by the high-copy- number mucA+B+ recombinant plasmid pGW1700 (27).…”

Section: Resultsmentioning

confidence: 95%

“…Both umuD and umuC mutants are virtually nonmutable with UV and many chemicals (5,9,36). The mucAB operon is a plasmid-borne analog of the umuDC operon (26,27,44) and is encoded by the 35.4-kb N-incompatibility group plasmid pKM101 (33,46). pKM101 is one of a number of extrachromosomal elements that enhance both the resistance of their hosts to DNA-damaging agents and their mutagenesis by these agents (37,42).…”

mentioning

confidence: 99%

“…TnS and Tn1000 insertion mutants of muc have been obtained in screens for plasmid derivatives that are unable to enhance base substitution mutagenesis (27,33). Each of these mutants is also unable to enhance resistance to UV killing.…”

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confidence: 99%

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LexA-independent expression of a mutant mucAB operon

1990

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pKM101 is a naturally occurring plasmid that carries mucAB, an analog of the umuDC operon, the gene products of which are required for the SOS-dependent processing of damaged DNA necessary for most mutagenesis. Genetic studies have indicated that mucAB expression is controlled by the SOS regulatory circuit, with LexA acting as a direct repressor. pGW16 is a pKMl01 derivative obtained by N-methyl-N'-nitro-Nnitrosoguanidine mutagenesis that was originally identified on the basis of its ability to cause a modest increase in spontaneous mutation rate. In this report, we show that pGW16 differs from pKM101 in being able to enhance methyl methanesulfonate mutagenesis and to confer substantial resistance to UV killing in a lexA3 host. The mutation carried by pGW16 is dominant and was localized to a 2.4-kb region of pGW16 that includes the mucAB coding region and approximately 0.6 kb of the 5'-flanking region. We determined the sequence of a 119-bp fragment containing the region upstream of mucAB and identified a single-base-pair change in'that region, a G. C-to-A T transition that alters a sequence homologous to known LexA-binding sites. DNA gel shift experiments indicate that LexA protein binds poorly to a 125-bp fragment containing this mutation, whereas a fragment containing the wild-type sequence is efficiently bound by LexA. This mutation also alters an overlapping sequence that is homologous to the -10 region of Escherichia coli promoters, moving it closer to the consensus sequence. The observation that the synthesis of pGW16-encoded mucAB proteins in maxicells is increased relative to that of pKM101-encoded mucAB proteins even in the absence of a lexA+ plasmid suggests that this mutation also increases the activity of the mucAB promoter.

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Section: Resultsmentioning

confidence: 95%

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confidence: 99%

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LexA-independent expression of a mutant mucAB operon

1990

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“…The study with S-9 mix used S. typhimurium TA98 carrying plasmid pKM101, which has higher sensitivity for DNA damage due to SOS repair response (37,38). On the contrary, S. typhimurium TA1538 and TA1538/pYG219 were used for detecting the HCAs mutagenicity in the chemical model since, speciˆcally, frame-shift type mutation was detected.…”

Section: Resultsmentioning

confidence: 99%

Mutagenicity of Heterocyclic Amines by Biomimetic Chemical Models for Cytochrome P450 in Ames Assay

Inami

Nagao

Ishikawa

et al. 2010

Genes and Environment

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Heterocyclic amines (HCAs) are a family of mutagenic and carcinogenic compounds produced during cooking or other burning processes, and exist in the environment. HCAs are metabolically activated by cytochrome P450, conjugated by phase II enzymes, to react with guanine bases. The aim of this study is to establish a chemical model for cytochrome P450 as an alternative to S-9 mix for detecting HCA mutagenicity in Salmonella strain. A chemical model was developed by comparing the mutagenicity of 3-amino-1-methyl-5H-pyrido [4,3-b]indole (Trp-P-2) in the presence of an iron porphyrin and an oxidant. The iron porphyrin derivatives, water-soluble 5,10,15,20-tetrakis(1-methylpyridinium-4-yl)porphyrinatoiron (III) chloride (4-MPy) or water-insoluble 5,10,15,20-tetrakis (penta‰uorophenyl)porphyrinatoiron (III) chloride (F 5 P), and the oxidant tert-butyl hydroperoxide (t-BuOOH), magnesium monoperoxyphthalate or iodosylbenzene were used. 4-MPy or F 5 P with t-BuOOH activated Trp-P-2, and the activity was similar with either porphyrin. Water-soluble model has a better chance to detect unstable compound, since the tester strain was exposed in the whole incubation period in the mutation procedure with 4-MPy. The eŠectiveness of 4-MPy/t-BuOOH was evaluated with other HCAs; IQ, MeIQ, MeIQx, Glu-P-1, Glu-P-2, PhIP, Trp-P-1, MeAaC and AaC. All HCAs except for MeAaC and AaC were mutagenic in Salmonella typhimurium TA1538. MeAaC and AaC were not mutagenic in TA1538, but they were mutagenic in S. typhimurium TA1538/pYG219, which overexpresses O-acetyltranferase on the TA1538 genetic background. Although the HCAs mutagenicity with the chemical model was weaker than that with S-9 mix, the chemical models activated HCAs without S-9 mix in the Ames assay.

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“…Starting material for construction of the plasmid was the plasmid pGW1700 (10), kindly provided by David Sobell of Graham Walker's laboratory (Massachusetts Institute of Technology, Cambridge), which contains the two muc genes from pKM101 but is considerably smaller (15). The genes were subcloned into a derivative of pBR322 (10).…”

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confidence: 99%

A plasmid carrying mucA and mucB genes from pKM101 in Haemophilus influenzae and Escherichia coli

Spikes

Setlow

1989

J Bacteriol

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The plasmid pMucAMucB, constructed from the Haemophilus influenzae vector pDM2, and a similar plasmid, constructed from pBR322, increased the survival after UV irradiation of Escherichia coli AB1157 with the umu-36 mutation and also caused UV-induced mutation in the E. coli strain. In H. influenzae, pMucAMucB caused a small but reproducible increase in survival after UV irradiation in wild-type cells and in a rec-l mutant, but there was no increase in spontaneous mutation in the wild type or in the rec-l mutant and no UV-induced mutation.We showed earlier (1) that a plasmid containing the mucA gene from pKM101 caused UV mutagenesis in Haemophilus influenzae, which is normally not operative in this species (4, 7). We now report on the biological effects in H. influenzae and Escherichia coli of a plasmid, pMucAMucB, we have constructed that carries both mucA and mucB genes from pKM101 in our cloning vector pDM2 (5).Starting material for construction of the plasmid was the plasmid pGW1700 (10), kindly provided by David Sobell of Graham Walker's laboratory (Massachusetts Institute of Technology, Cambridge), which contains the two muc genes from pKM101 but is considerably smaller (15). The genes were subcloned into a derivative of pBR322 (10). A 2.4-kilobase-pair fragment was cut out of pGW1700 with the restriction endonucleases XmnI and Scal, and PstI linkers were attached, making it possible to clone the fragment in the vector pDM2, which has a unique PstI site (5), and also to remove the fragment from pDM2. The fragment with PstI linkers was first inserted into PstI-cut pBR322 in order to amplify the PstI-ended fragment for successful cloning in H. influenzae via pDM2. Restriction endonuclease mapping of the plasmid showed the presence of both muc genes, as judged by comparison with previous mapping of these genes (9). H. influenzae and E. coli were transformed by the plasmids as previously described (11,17). Crude plasmid preparations (8) were used to transfer pMucAMucB between H. influenzae species. Figure 1 shows survival curves of AB1157 umu-36 (GW3198), kindly supplied by Graham Walker and previously described (3), containing plasmids pDM2 and pBR322 with and without the two muc genes. The inserts increased the survival to the same extent, whether in pDM2 or pBR322. Thus, it was clear that the lethal effect of UV irradiation of E. coli was similarly influenced by the muc genes in the two different plasmids. The survival was not affected by the plasmids pDM2 and pBR322. The data are in accord with the increase in survival seen in E. coli strain TK610 (AB1157 umuC36 and also containing a mutation affecting excision of pyrimidine dimers) with the plasmid pSK100 carrying both the umuD and umuC genes (16). Figure 2 shows survival curves of wild-type strain Rd with the cloning vector pDM2 and with the vector containing the muc genes. The strains were irradiated together to eliminate possible differences in dosimetry and were distinguished on plates by appropriate antibiotics in the agar. The difference in UV sensi...

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Localization of the plasmid (pKM101) gene(s) involved in recA + lexA +-dependent mutagenesis

Cited by 111 publications

References 31 publications

LexA-independent expression of a mutant mucAB operon

LexA-independent expression of a mutant mucAB operon

Mutagenicity of Heterocyclic Amines by Biomimetic Chemical Models for Cytochrome P450 in Ames Assay

A plasmid carrying mucA and mucB genes from pKM101 in Haemophilus influenzae and Escherichia coli

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