Biomedical subjects

T Nohmi

Publications and source records attributed to T Nohmi.

At least 55 records · Page 3Linked to original sources

PubMed1998

Establishment of a Salmonella tester strain highly sensitive to mutagenic heterocyclic amines.

Heterocyclic amines (HCAs) that are present in cooked foods require metabolic activation to exert their genotoxicity. They undergo activation via N-hydroxylation by cytochrome P450 1A2 (CYP1A2), followed by O-esterification by O-acetyltransferase (OAT). To develop a Salmonella tester strain that is highly sensitive to mutagenic HCAs, we introduced a coexpression plasmid (p1A2OR) carrying human CYP1A2 and NADPH-CYP reductase cDNAs and an expression plasmid (pOAT) carrying Salmonella OAT to Salmonella typhimurium TA1538 to yield a TA1538/ARO strain. The TA1538/ARO strain was proven to express the enzymes, as indicated by high activities of 7-ethoxyresorufin O-deethylase and isoniazid N-acetylase. The TA1538/ARO strain exhibited very high sensitivity to mutagenic HCAs 2-amino-3,4-dimethylimidazo[4,5-f]quinoline, 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), and 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline and a somewhat higher sensitivity to 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine compared with the parent Ames tester strain TA1538. The minimum concentrations of 2-amino-3,4-dimethylimidazo[4,5-f]quinoline, IQ, 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline, and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine giving positive results were defined by evidence that the number of colonies increased in a dose-dependent manner and reached a number two times higher than that obtained by vehicle alone as a control in the TA1538/ARO strain at concentrations of 0.3, 3, 30, and 1000 pM, respectively. When the membrane and cytosol fractions prepared from TA1538/ARO were added to a mixture containing the parental TA1538, the sensitivity of TA1538 to IQ was much lower than that seen with TA1538/ARO. These results indicate that the intracellular expression of drug-metabolizing enzymes makes the established strain of Salmonella highly sensitive to mutagenic HCAs.

Acetyltransferases↗

PubMed1998

Roles of the mutagenesis proteins SamA'B and MucA'B in chemically induced frameshift mutagenesis in Salmonella typhimurium hisD3052.

The mutagenesis induced by ultraviolet light and many chemicals in Escherichia coli is largely dependent upon the proteins encoded by the umuDC operon and their analogs. In Salmonella typhimurium, there are two sets of umuDC-like operons: the umuDC(ST) operon in the chromosome and the samAB operon located on the 60-MDa cryptic plasmid. The former operon, but not the latter, confers UV mutability on S. typhimurium. Nevertheless, the samAB operon, when carried on high-copy-number plasmids, can efficiently promote UV mutagenesis. In order to characterize the function of samAB in greater detail, we have compared the abilities of MucA'B and a putative activated form of SamAB, i.e. SamA'B, to promote chemically induced frameshift mutagenesis in S. typhimurium hisD3052. MucAB is an activated form of the products of mucAB, which is the most potent umuDC analog characterized so far. We have used four plasmids, each carrying samA', samB, mucA' or mucB with a lac promoter instead of their own promoters. The results indicated that under the conditions of elevated expression, SamA'B can promote chemically induced frameshift mutagenesis by furylfuramide, aflatoxin B1, 1-nitropyrene, and 1,8-dinitropyrene, with efficiencies comparable to, or even better than, MucA'B. Increase of the levels of expression enhanced the ability of SamA'B to promote the mutagenesis, while it decreased that of MucA'B. Surprisingly, the elevated expression of MucB alone significantly enhanced the frameshift mutagenesis induced by 1-nitropyrene and 1,8-dinitropyrene, whereas the elevated expression of SamB, MucA' and SamA' did not enhance it. These results suggest that the abilities of SamA'B and MucA'B to promote mutagenesis strongly depend on their levels of expression. The possible roles of these mutagenesis proteins in chemically induced frameshift mutagenesis are discussed.

Bacterial Proteins↗

PubMed1998

Genomic structure and chromosomal localization of the mouse Ogg1 gene that is involved in the repair of 8-hydroxyguanine in DNA damage.

8-Hydroxyguanine (7,8-dihydro-8-oxoguanine: oh8Gua) is a damaged form of guanine induced by oxygen-free radicals and causes GC to TA transversions. Previously we isolated the hOGG1 gene, a human homolog of the yeast OGG1 gene, which encodes a DNA glycosylase and lyase to excise oh8Gua in DNA. In this study, we isolated a mouse homolog (Ogg1) of the OGG1 gene, characterized oh8Gua-specific DNA glycosylase/AP lyase activities of its product, and determined chromosomal localization and exon-intron organization of this gene. A predicted protein possessed five domains homologous to human and yeast OGG1 proteins. Helix-hairpin-helix and C2H2 zinc finger-like DNA-binding motifs found in human and yeast OGG1 proteins were also retained in mouse Ogg1 protein. The properties of a GST fusion protein were identical to human and yeast OGG1 proteins in glycosylase/lyase activities, their substrate specificities, and suppressive activities against the spontaneous mutagenesis of an Escherichia coli mutM mutY double mutant. The mouse Ogg1 gene was mapped to Chromosome (Chr) 6, and consisted of 7 exons approximately 6 kb long. Two DNA-binding motifs were encoded in exons 4 through 5. These data will facilitate the investigation of the OGG1 gene to elucidate the relationship between oxidative DNA damage and carcinogenesis.

Amino Acid Sequence↗

PubMed1997

Multiple pathways for SOS-induced mutagenesis in Escherichia coli: an overexpression of dinB/dinP results in strongly enhancing mutagenesis in the absence of any exogenous treatment to damage DNA.

dinP is an Escherichia coli gene recently identified at 5.5 min of the genetic map, whose product shows a similarity in amino acid sequence to the E. coli UmuC protein involved in DNA damage-induced mutagenesis. In this paper we show that the gene is identical to dinB, an SOS gene previously localized near the lac locus at 8 min, the function of which was shown to be required for mutagenesis of nonirradiated lambda phage infecting UV-preirradiated bacterial cells (termed lambdaUTM for lambda untargeted mutagenesis). A newly constructed dinP null mutant exhibited the same defect for lambdaUTM as observed previously with a dinB::Mu mutant, and the defect was complemented by plasmids carrying dinP as the only intact bacterial gene. Furthermore, merely increasing the dinP gene expression, without UV irradiation or any other DNA-damaging treatment, resulted in a strong enhancement of mutagenesis in F'lac plasmids; at most, 800-fold increase in the G6-to-G5 change. The enhanced mutagenesis did not depend on recA, uvrA, or umuDC. Thus, our results establish that E. coli has at least two distinct pathways for SOS-induced mutagenesis: one dependent on umuDC and the other on dinB/P.

Bacterial Proteins↗

PubMed1997

New tester strains of Salmonella typhimurium lacking O6-methylguanine DNA methyltransferases and highly sensitive to mutagenic alkylating agents.

Salmonella typhimurium YG7104 and YG7108 are derivatives of the Ames tester strain TA1535, and have chromosomal deletions of the ogtST gene or both the ogtST and adaST genes, respectively. The ogtST and adaST genes encode O6-methylguanine DNA methyltransferases that are involved in the repair of DNA damage caused by alkylating agents. The sensitivities of these strains to 15 mutagens with different structures were tested and compared with those of the parent strain TA1535. Deletion of ogtST or ogtST plus adaST substantially increased the sensitivity of strain TA1535 to the mutagenicity of alkylating agents, such as N-ethyl-N'-nitro-N-nitrosoguanidine, ethyl methanesulfonate or dimethylnitrosamine (DMN). Preincubation of the chemical with S9 mix and bacteria for 20 min at 37 degrees C before pouring them together on agar plates was not necessary to detect the mutagenicity of DMN when strain YG7104 or YG7108 was used as a tester strain. Introduction of plasmid pKM101 did not enhance but rather decreased the sensitivity of YG7104 and YG7108 to alkylating agents. Since the new strains are highly sensitive only to alkylating agents, they will be useful to detect the mutagenicity with high efficiency and to study the mechanism of mutagenesis induced by environmental alkylating agents.

Alkylating Agents↗

PubMed1997

Construction of mutants of Salmonella typhimurium deficient in 8-hydroxyguanine DNA glycosylase and their sensitivities to oxidative mutagens and nitro compounds.

8-Hydroxyguanine (8-OH-G) DNA glycosylase is an enzyme involved in repair of oxidative DNA damage, e.g., 8-OH-G in DNA. In order to assess the roles of 8-OH-G in spontaneous and chemically-induced mutagenesis, the mutMST gene encoding 8-OH-G DNA glycosylase of Salmonella typhimurium was disrupted in several Ames tester strains, i.e., S. typhimurium TA1535 (hisG46, uvrB-, rfa), TA1975 (hisG46, uvr+, rfa) and TA102 (hisG428, uvr+, rfa). The spontaneous mutation frequencies were increased 2.4 and 1.6 times, respectively, by the mutMST deletions in strains TA1535 and TA1975, which are spontaneously reverted to His+ by mutations mainly at G:C base pairs. The resulting strains YG3001 (TA1535 delta mutMST) and YG3002 (TA1975 delta mutMST) were 2 to 8 times more sensitive to the mutagenicities of methylene blue plus visible light, neutral red plus visible light and 2-nitrofluorene than the parent strains. The strain YG3002 but not YG3001 was about 30 times more sensitive to the mutagenicity of 4-nitroquinoline N-oxide than the parent strain TA1975. Neither hydrogen peroxide nor phenazine methosulfate was mutagenic in the mutMST-deletion strains as well as in the parent strains. In contrast, the mutMST deletion did not affect the spontaneous mutation frequency of strain TA102, which has an A:T base pair at the critical site for reversion. The sensitivities of strain TA102 to the chemicals were not enhanced by the mutMST deletion except for hydrogen peroxide. These results suggest that 8-OH-G in DNA plays important roles in spontaneous mutagenesis occurring at G:C base pairs in S. typhimurium, and some nitro aromatics such as 4-nitroquinoline N-oxide or 2-nitrofluorene as well as the photosensitizers plus visible light can produce 8-OH-G in DNA, thereby inducing mutations. In the case of 4-nitroquinoline N-oxide, 8-OH-G rather than DNA adducts seems to play major roles in mutagenesis in uvr+ background. The new strains could be useful for the evaluation of the roles of 8-OH-G in mutagenesis in S. typhimurium and permit the efficient detection of some oxidative mutagens in the environment.

4-Nitroquinoline-1-oxide↗

PubMed1997

Cloning of a human homolog of the yeast OGG1 gene that is involved in the repair of oxidative DNA damage.

We report the cloning of a human homolog of the yeast OGGC1 gene, which encodes a DNA glycosylase that excises an oxidatively damaged form of guanine, 8-hydroxyguanine (also known as 7,8-dihydro-8-oxoguanine). Since the deduced amino acid sequence (68 amino acids) of a human expressed sequence tag, N55394, matched a short stretch of yeast OGG1 protein with greater than 40% amino acid identity, a full length cDNA clone was isolated from a HeLa cell cDNA library with the N55394 clone as a probe. The cDNA clone encodes a predicted protein of 345 amino acids which is homologous to yeast OGG1 protein throughout the entire polypeptide sequence and shares 38% amino acid identity with yeast OGG1 protein. Moreover, we found that both a human homolog and yeast OGG1 protein possess two distinct DNA binding motifs, a helix-hairpin-helix (HhH) motif and a C2H2 zinc finger like motif, and a domain homologous to human and E. coli MutY proteins. Expression of a human homolog suppressed spontaneous mutagenesis of an E. coli (mutM mutY) mutant as in the case of yeast OGG1 protein. The gene was ubiquitously expressed in a variety of human organs and mapped to chromosome 3p26.2. These results strongly suggest that the gene isolated here is a human counterpart of the yeast OGGI gene and is involved in the repair of oxidative DNA damage in human cells.

Amino Acid Sequence↗

PubMed1997

Targeted disruption of the gene encoding the classical nitroreductase enzyme in Salmonella typhimurium Ames test strains TA1535 and TA1538.

The gene encoding the 'classical nitroreductase' (CNR) of Salmonella typhimurium was disrupted. In this manner, cnr null mutant derivatives of strains TA1535 and TA1538 were constructed, and named YG7131 and YG7127, respectively. In both strain backgrounds, cnr gene disruption reduced nitrofurazone-reductase activity. This reduction almost completely eliminated the nitroreductase activity of strain TA1538. In contrast, the nitroreductase activity of strain TA1535 was much higher than that in TA1538. In this background, cnr gene disruption resulted in a reduction in nitroreductase activity by a similar absolute amount as in TA1538, but representing only about one-quarter of the original activity of TA1535. The results suggest that S. typhimurium has originally at least two distinct nitroreductases, one of which is already deficient in strain TA1538; the CNR is present in both TA1535 and TA1538. Also, these two strains (including their derivatives, TA98 and TA100) are not isogenic with regard to nitroreductase activity. After the introduction of plasmid pKM101, the sensitivities of the strains YG7132 and YG7128, the cnr-null mutants of TA98 and TA100, respectively, against several nitro compounds were compared with those of the conventional cnr-deficient strains TA98NR and TA100NR and the wild-type strains TA98 and TA100. The mutagenicities of 2-nitrofluorene and 1-nitropyrene in YG7132 or TA98NR were ten-fold lower than those of the compounds in TA98. Similarly, the mutagenicity of 2-(2-furyl)-3-(5-nitro-2-furyl) acrylamide in strain YG7128 or TA100NR was substantially lower than that of the compound in TA100. However, the mutagenicity of 2-nitronaphthalene in YG7128 was between those observed with TA100 and TA100NR, suggesting that a nitroreductase in S. typhimurium other than CNR is involved in the metabolic activation of this compound. The cnr gene of S. typhimurium positively hybridized with DNA at 13 min on the E. coli chromosome where the nfsB and nfnB genes of E. coli are mapped. These results suggest that the cnr gene of S. typhimurium is a counterpart of the nfsB and nfnB genes of E. coli, and that the newly constructed cnr-deletion strains are useful to assess the role of nitroreductases in the metabolic activation of mutagenic nitro compounds.

Blotting, Southern↗

PubMed1997

Recent advances in the construction of bacterial genotoxicity assays.

Bacterial mutagenicity assays have been widely used in genotoxicology research for two decades. We discuss the development of such assays, especially the Ames test, with particular attention to strain engineering. Genes encoding enzymes of mutagen bioactivation, including N-acetyltransferase, nitroreductase, and cytochrome P450, have been introduced into tester strains. The processing of DNA damage by the bacterial strains has also been modified in several ways, so as to enhance mutagenesis. These efforts have greatly increased the sensitivity of mutation assays and have illuminated the molecular mechanisms of mutagenesis. We also discuss the relationship between bacterial assays and in vivo mutation assays which use transgenic rodents.

Air Pollutants↗

PubMed1996

Construction of a new system for separate expression of mutagenesis proteins: the abilities to promote UV mutagenesis and interchangeability of MucA', MucB, SamA' and SamB proteins in Salmonella typhimurium.

The two distinct mucAB and samAB operons originally isolated from the plasmids of Salmonella typhimurium encode proteins engaged in induced mutagenesis. They represent two extreme cases among the so far characterized members of the enterobacterial umuDC family in respect to both the strength and the specificity of their effect. It is suggested that the MucA and SamA proteins are post-translationally processed to MucA' and SamA', respectively, which lack the N-terminal 25 amino acids and are the active species in mutagenesis. For the purpose of characterizing the individual activities of these proteins, we developed a new system for their SOS-independent separate and controllable expression in enterobacteria. Besides the matured forms of MucA', SamA' as well as MucB and SamB proteins we also expressed hybrid HisTag-MucA' and HisTag-SamA' proteins in which a synthetic 24 amino acid HisTag region replaces the natural 25 amino acid N-terminal leader present in the MucA and SamA precursors. In this study, we analyzed the effect of the mutagenesis proteins on the UV mutability of S. typhimurium YG5144. None of the proteins, if expressed alone, promoted UV mutagenesis. Different combinations of the proteins promoted mutagenesis to different extents in the order MucA' + MucB > SamA' + SamB > or = HisTag-MucA' + MucB > or = SamA' + MucB > MucA' + SamB > HisTag-SamA' + SamB. The mutagenesis enhancing potential of the combinations with MucB protein decreased as the expression of the proteins increased while the mutagenesis enhancing potential of the combinations with SamB protein increased together with the increase in the expression. The artificially expressed MucA' + MucB proteins were as active as their MucAB counterparts expressed from the plasmid pKM101 in promoting UV mutagenesis, but they were remarkably more efficient than their pKM101-born counterparts in promoting spontaneous mutagenesis. We conclude that the MucA'B and SamA'B proteins are partly interchangeable and the functionality of the resulting A' + B complex is largely dependent on the appropriate B-protein.

Amino Acid Sequence↗

PubMed1996

A new transgenic mouse mutagenesis test system using Spi- and 6-thioguanine selections.

A new transgenic mouse mutagenesis test system has been developed for the efficient detection of point mutations and deletion mutations in vivo. The mice carry lambda EG10 DNA as a transgene. When the rescued phages are infected into Escherichia coli YG6020-expressing Cre recombinase, the phage DNA is converted into plasmid pYG142 carrying the chloramphenicol-resistance gene and the gpt gene of E. coli. The gpt mutants can be positively detected as colonies arising on plates containing chloramphenicol and 6-thioguanine. The EG10 DNA carries a chi site along with the red and gam genes so that the wild-type phages display Spi- (sensitive to P2 interference) phenotype. Mutant phages lacking both red and gam genes can be positively detected as plaques that grow in P2 lysogens of E. coli. These mutant phages are called lambda Spi-. The spontaneous gpt mutation frequencies of five independent transgenic lines were 1.7 to 3.3 x 10(-5) in bone marrow. When the mice were treated with ethylnitrosourea (single i.p. treatments with 150 mg/kg body weight; killed 7 days after the treatments), mutation frequencies were increased four- to sevenfold over the background in bone marrow. The average rescue efficiencies were more than 200,000 chloramphenicol-resistant colonies per 7.5 micrograms bone marrow DNA per packaging reaction. In contrast to gpt mutation frequencies, spontaneous Spi- mutation frequencies were 1.4 x 10(-6) and 1.1 x 10(-6) in bone marrow and sperm, respectively. No spontaneous Spi- mutants have been detected so far in spleen, although 930,000 phages rescued from untreated mice were screened. In gamma-ray-treated animals, however, induction of Spi- mutations was clearly observed in spleen, at frequencies of 1.4 x 10(-5) (5 Gy), 1.2 x 10(-5) (10 Gy), and 2.0 x 10(-5) (5O Gy). These results suggest that the new transgenic mouse "gpt delta" could be useful for the efficient detection of point mutations and deletion mutations in vivo.

Animals↗

PubMed1996

Regionally-targeted mutagenesis by metabolically-activated steviol: DNA sequence analysis of steviol-induced mutants of guanine phosphoribosyltransferase (gpt) gene of Salmonella typhimurium TM677.

Steviol is the aglycone of stevioside, a non-caloric sugar substitute commonly used in Japan. Steviol strongly induces mutations at the guanine phosphoribosyltransferase gene (gpt) of Salmonella typhimurium TM677 when the metabolic activation system (S9 mix) is present. However, it is completely negative in the reverse mutation assays using Escherichia coli WP2uvrA/pKM101 or S.typhimurium TA strains. In order to characterize the mutations induced by metabolically-activated steviol, the chromosomal gpt alleles of 24 induced (ST clones) and 16 spontaneous mutants (SP clones) of S.typhimurium TM677 were sequenced and the mutation spectra were compared. About 40% of the mutations of ST clones (nine out of 24) were localized in the region between nucleotides 280 and 330 from the starting codon ATG, whereas no mutations of SP clones were found in that region. The mutations identified in the region included transitions (three clones), transversions (four clones), a duplication and a deletion. There were no other marked differences between ST and SP clones: base-change mutations were dominant over frameshifts and deletions (ST clones, 20 versus three; SP clones, 16 versus two) and base change mutations occurred more frequently at G:C pairs rather than at A:T pairs (ST clones, 15 versus five; SP clones, 12 versus four). The possibility that metabolically-activated steviol gives pause to DNA synthesis around nucleotide 280, thereby stimulating the duplication, deletion and untargeted mutagenesis in the defined region of the gpt gene is discussed.

Amino Acid Sequence↗

PubMed1996

Evaluation of the genotoxicity of stevioside and steviol using six in vitro and one in vivo mutagenicity assays.

Stevioside, a constituent of Stevia rebaudiana, is commonly used as a non-caloric sugar substitute in Japan. The genetic toxicities of stevioside and its aglycone, steviol, were examined with seven mutagenicity tests using bacteria (reverse mutation assay, forward mutation assay, umu test and rec assay), cultured mammalian cells (chromosomal aberration test and gene mutation assay) and mice (micronucleus test). Stevioside was not mutagenic in any of the assays examined. The aglycone, steviol, however, produced dose-related positive responses in some mutagenicity tests, i.e. the forward mutation assay using Salmonella typhimurium TM677, the chromosomal aberration test using Chinese hamster lung fibroblast cell line (CHL) and the gene mutation assay using CHL. Metabolic activation systems containing 9000 g supernatant fraction (S9) of liver homogenates prepared from polychlorinated biphenyl or phenobarbital plus 5,6-benzoflavone-pretreated rats were required for mutagenesis and clastogenesis. Steviol was weakly positive in the umu test using S.typhimurium TA1535/pSK1002 either with or without the metabolic activation system. Steviol, even in the presence of the S9 activation system, was negative in other assays, i.e. the reverse mutation assays using S.typhimurium TA97, TA98, TA100, TA102, TA104, TA1535, TA1537 and Escherichia coli WP2 uvrA/pKM101 and the rec-assay using Bacillus subtilis. Steviol was negative in the mouse micronucleus test. The genotoxic risk of steviol to humans is discussed.

Animals↗

PubMed1996

Prophylactic effect of Enterococcus faecalis FK-23 preparation on experimental candidiasis in mice.

The prophylactic effects of heat-killed cells of Enterococcus faecalis FK-23 (FK-23 preparation) on experimental candidiasis were investigated in normal and leukopenic mice. In cyclophosphamide-induced leukopenic mice, oral or intraperitoneal administration of the FK-23 preparation at a daily dose of 1.25 or 5 mg/mouse for 3 consecutive days prior to Candida albicans infection significantly prolonged survival periods of the infected mice, and decreased viable counts of C. albicans recovered from their kidneys. In normal mice, the FK-23 preparation administered at dosages ranging from 0.63 to 10 mg/mouse/day for 3 consecutive days was ineffective, while in leukopenic mice, the FK-23 administered orally caused a facilitated recovery in the number of white blood cells including neutrophils. Furthermore, intraperitoneal administration of the FK-23 preparation into mice augmented the anti-Candida activity of immunocompromised peritoneal exudate cells obtained from the animals. These results suggested the potential usefulness of the FK-23 preparation as a prophylactic agent for the management of patients with opportunistic fungal infections.

Animals↗

PubMed1996

A glucocorticoid antagonist, mifepristone affects anti-Candida activity of murine neutrophils in the presence of prednisolone in vitro and experimental candidiasis of prednisolone-treated mice in vivo.

The effects of a glucocorticoid-antagonist, mifepristone on the suppressive action of prednisolone for anti-Candida activity of murine neutrophils were examined. Prednisolone suppressed inhibitory activity of neutrophils to mycelial growth of Candida albicans. This suppression was cancelled in the presence of 10(-7)-10(-6) M of mifepristone in vitro. Corresponding to this in vitro action, mifepristone protected prednisolone-treated mice from lethal C. albicans infection in vivo. These results suggest that glucocorticoid-induced vulnerability to Candida infection may be recovered or normalized by application of mifepristone.

Animals↗

PubMed1996

[Augmented ability of spleen cells to produce interferons and prevention from lethal infection of herpes simplex virus in mice orally treated with Enterococcus faecalis preparation, FK-23].

Effects of the oral or intraperitoneal administration of an Enterococcus preparation, FK-23, to mice on the interferon (IFN) production by their spleen cells and on the host defense against the infection with herpes simplex virus (HSV)-1 were examined. Spleen cells were obtained from the mice intraperitoneally treated with cyclophosphamide (CY) and subsequently orally administered FK-23 preparation, and then cultured with phytohemagglutinin-P or bacterial lipopolysaccharide in vitro. They produced higher titers IFN than those obtained from control mice which were not treated with the FK-23 preparation. The IFN activity was neutralized mainly by antiIFN-beta antibody. Correspondingly, oral (5 mg/mouse) or intraperitoneal (1 mg/mouse) administration of the FK-23 preparation protected some of the CY-pretreated mice from death by HSV-1 infection.

Administration, Oral↗

PubMed1995

Involvement of umuDCST genes in nitropyrene-induced -CG frameshift mutagenesis at the repetitive CG sequence in the hisD3052 allele of Salmonella typhimurium.

Expression of the umuDC operon is required for UV and most chemical mutagenesis in Escherichia coli. The closely related species Salmonella typhimurium has two sets of umuDC-like operons, umuDCST on the chromosome and samAB on a 60-MDa cryptic plasmid. The roles of the umuDC-like operons in chemically induced frameshift mutagenesis of the hisD3052 allele of S. typhimurium were investigated. Introduction of a pBR322-derived plasmid carrying umuDCST increased the rate of reversion of hisD3052, following treatment with 1-nitropyrene (1-NP) or 1,8-dinitropyrene (1,8-DNP) tenfold and fivefold, respectively, whereas it did not substantially increase the rate of reversion induced by other frameshift mutagens, i.e. 2-nitrofluorene (2-NF) and 2-amino-3-methyldipyrido[1,2-a:3',2'-d]imidazole (Glu-P-1). Introduction of a pBR322-derived plasmid carrying samAB did not increase the incidence of reversion of hisD3052 observed with any of the mutagens examined. Deletion of umuDCST substantially lowered the reversion rate induced by 1-NP or 1,8-DNP, but it did not affect reversion induced by 2-NF, Glu-P-1 or N-hydroxyacetylaminofluorene (N-OH-AAF). Deletion of samAB had little impact on reversion incidence induced by any of the five frameshift mutagens. DNA amplification using the polymerase chain reaction technique followed by restriction enzyme analysis using BssHII, suggested that the mutations induced by the five frameshift mutagens were all CG deletions at the CGCGCGCG sequence in hisD3052. These results suggest that umuDCST, but not samAB, is involved in the -2 frameshift mutagenesis induced by 1-NP and 1,8-DNP at the repetitive CG sequence, whereas neither operon participates in induction of the same type of mutations by 2-NF, Glu-P-1 or N-OH-AAF.

Alcohol Oxidoreductases↗

PubMed1995

Specific disruption of samAB genes in a 60-megadalton cryptic plasmid of Salmonella typhimurium.

The expression of umuDC operon is required for UV and most chemical mutagenesis in Escherichia coli. The closely related species Salmonella typhimurium has two sets of umuDC-like operons, i.e., umuDCST in the chromosome and samAB in a 60-megadalton cryptic plasmid. In this study, we specifically disrupted the samAB genes to investigate their exact roles in UV mutagenesis in S. typhimurium. The specific gene disruption was carried out by the preligation method. Deletion of samAB did not lower the UV mutability of S. typhimurium TA2659 but rather increased the UV mutability about twofold. The samAB-umuDCST double deletion mutant as well as the umuDCST deletion mutant was UV nonmutable. These results suggest that the samAB genes do not considerably contribute to the UV mutability of S. typhimurium and raise the question of why such quiet umuDC-like genes are present in the cryptic plasmid of S. typhimurium.

Blotting, Southern↗