Of the three isomers of the aromatic amine phenylenediamine (PDA), only o-PDA, but not m- and p-PDA, induced DNA damage (as measured by the Comet assay), and somatic mutations in the leaves of the chlorophyll-deficient tester strain Nicotiana tabacum var. xanthi. With increasing light intensity (0, 30, 80 or 140 micromol m(-2)s(-1) photosynthetic photon fluence rate) during a 72h mutagenic treatment of tobacco seedlings, o-PDA-induced DNA damage and the yield of somatic mutations were significantly increased. The peroxidase inhibitor diethyldithiocarbamate (DEDTC) repressed o-PDA-induced DNA damage. The effect of light is caused by the light-dependent increase of peroxidase activity and the accumulation of hydrogen peroxide, which participate in the metabolic activation of the promutagen o-PDA to mutagenic product(s). In contrast, DNA damage induced by the direct-acting alkylating mutagen ethyl methanesulphonate was the same whether treatment was in the light or in the dark, and was not repressed by the peroxidase inhibitor DEDTC.

• MeSH
• chlorofyl nedostatek   genetika   MeSH
• ethylmethansulfonát toxicita   MeSH
• fenylendiaminy metabolismus   toxicita   MeSH
• jedovaté rostliny * MeSH
• kometový test MeSH
• listy rostlin účinky léků   enzymologie   MeSH
• mutace * MeSH
• mutageny metabolismus   toxicita   MeSH
• peroxid vodíku metabolismus   MeSH
• peroxidasa metabolismus   MeSH
• poškození DNA * MeSH
• světlo škodlivé účinky   MeSH
• tabák účinky léků   enzymologie   genetika   účinky záření   MeSH
• vztah dávky záření a odpovědi MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• 1,2-diaminobenzene MeSH Prohlížeč
• chlorofyl MeSH
• ethylmethansulfonát MeSH
• fenylendiaminy MeSH
• mutageny MeSH
• peroxid vodíku MeSH
• peroxidasa MeSH

The advantage of using the tobacco (Nicotiana tabacum var. xanthi) mutagenicity assay is the ability to analyze and compare on the same plants under identical treatment conditions both the induced acute DNA damage in somatic cells as measured by the Comet assay and the yield of induced leaf somatic mutations. Gamma-irradiation of tobacco seedlings induced a dose-dependent increase in somatic mutations from 0.5 (control) to 240 per leaf (10Gy). The increased yield of somatic mutations was highly correlated (r = 0.996) with the increased DNA damage measured by the Comet assay immediately after irradiation. With increased dose of gamma-irradiation, the averaged median tail moment values ( +/- S.E.) significantly increased from 1.08 +/- 0.10 (control) to 20.26 +/- 1.61 microm (10Gy). Nuclei isolated from leaves 24h after irradiation expressed tail moment values that were not significantly different from the control (2.08 +/- 0.11). Thus a complete repair of DNA damage induced by gamma-irradiation and measurable by the Comet assay was observed, whereas the yield of somatic mutations increased in relation to the radiation dose. Data on the kinetics of DNA repair and of DNA damage induced by gamma-radiation on isolated tobacco nuclei, and on nuclei isolated from irradiated leaves and roots are presented.

• MeSH
• jedovaté rostliny * MeSH
• kometový test MeSH
• mutace * MeSH
• oprava DNA * MeSH
• poškození DNA * MeSH
• tabák genetika   účinky záření   MeSH
• záření gama MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

We employed single cell gel electrophoresis to analyze the kinetics of DNA repair in nuclei isolated from tobacco plants exposed to ethyl methanesulfonate (EMS), N-ethyl-N-nitrosourea (ENU) and gamma-radiation. DNA repair was measured as the reduction of the tail moment values as a function of time after the mutagen treatment ended. DNA damage in leaf nuclei of EMS-or ENU-treated tobacco plants persisted over a 72h recovery period. However, a reduction of the SCGE tail moment values in nuclei isolated from leaves was observed over a 4-week period of recovery. Newly emerged leaves expressed a lower level of DNA damage due to more efficient repair and/or dilution of initial DNA lesions during cell division. After 24h recovery, leaf nuclei from cells exposed to 20 or 40Gy of gamma-radiation expressed complete DNA repair. These data indicate that DNA lesions induced by alkylating agents are not readily repaired and persist beyond 4 weeks. Enzymes necessary to repair gamma-induced DNA lesions are fully functional in non-replicating leaf cells and single and double strand breaks are rapidly repaired.

• MeSH
• alkylační látky toxicita   MeSH
• buněčné jádro MeSH
• DNA účinky léků   účinky záření   MeSH
• elektroforéza MeSH
• ethylmethansulfonát toxicita   MeSH
• ethylnitrosomočovina toxicita   MeSH
• jedovaté rostliny * MeSH
• kometový test MeSH
• kořeny rostlin účinky léků   účinky záření   MeSH
• listy rostlin účinky léků   účinky záření   MeSH
• mutageny toxicita   MeSH
• oprava DNA * MeSH
• tabák účinky léků   genetika   účinky záření   MeSH
• záření gama * MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• alkylační látky MeSH
• DNA MeSH
• ethylmethansulfonát MeSH
• ethylnitrosomočovina MeSH
• mutageny MeSH

The plant growth regulator and herbicide maleic hydrazide (MH) induced a high frequency of somatic mutations in leaves of tobacco (Nicotiana tabacum var. xanthi) and a high yield of chromosome aberrations in roots of field beans (Vicia faba, karyotype ACB). In contrast, no significant increase in MH-induced DNA damage, as measured by the Comet assay, could be demonstrated in either plant species. The absence of DNA migration induced by MH was not effected in tobacco by either pH of the MH solution, the sampling time after MH treatment or continuous MH treatment for 14 days. To our knowledge, MH represents the first agent which has proved to be highly mutagenic and clastogenic but does not cause DNA damage as measured by the Comet assay in the same experimental system.

• MeSH
• časové faktory MeSH
• chromatidy účinky léků   ultrastruktura   MeSH
• chromozomální aberace MeSH
• DNA rostlinná účinky léků   MeSH
• Fabaceae účinky léků   genetika   růst a vývoj   MeSH
• herbicidy toxicita   MeSH
• hydrazid kyseliny maleinové toxicita   MeSH
• jedovaté rostliny * MeSH
• kometový test * MeSH
• koncentrace vodíkových iontů MeSH
• kořeny rostlin genetika   MeSH
• léčivé rostliny * MeSH
• listy rostlin genetika   MeSH
• mutace * MeSH
• mutageneze MeSH
• poškození DNA * MeSH
• tabák účinky léků   genetika   růst a vývoj   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• DNA rostlinná MeSH
• herbicidy MeSH
• hydrazid kyseliny maleinové MeSH

The use of single cell gel electrophoresis (SCGE) has recently been applied to plant systems. We optimized the experimental conditions for SCGE analysis using nuclei isolated from different tissues of intact plants. Concentration-response curves of genomic DNA migration were analyzed in intact plants treated with the monofunctional alkylating agents ethyl methanesulfonate (EMS), methyl methanesulfonate (MMS), N-ethyl-N-nitrosourea (ENU), and N-methyl-N-nitrosourea (MNU). These data were used to calibrate SCGE tail moment values to induced somatic mutation in plant leaves. We used a genotoxicity index to compare genomic DNA damage and the induction of somatic mutation in the leaf tissues. The rank order of the genotoxic potency of these alkylating agents assayed by SCGE was MNU >> MMS > ENU > EMS. The rank order for the mutagenic potency of these agents was MNU >> ENU congruent with MMS > EMS. The data demonstrate the utility of SCGE analysis in plant systems. The use of SCGE will permit a larger range of plants for use as in situ environmental monitors. Also, this approach may be used to search for crop plant germplasm accessions with enhanced genomic stability. We investigated whether the intragenomic distributions of DNA damage induced by these alkylating agents were uniform and random. When a plot of the ratio of the %tail DNA and tail length versus the concentration of the test mutagen was generated, the induced SCGE data deviated from a random distribution of genomic DNA damage.

• MeSH
• alkylační látky toxicita   MeSH
• DNA rostlinná chemie   účinky léků   MeSH
• elektroforéza metody   MeSH
• ethylmethansulfonát toxicita   MeSH
• ethylnitrosomočovina toxicita   MeSH
• jedovaté rostliny MeSH
• konformace nukleové kyseliny MeSH
• kořeny rostlin účinky léků   genetika   MeSH
• listy rostlin účinky léků   genetika   MeSH
• methylmethansulfonát toxicita   MeSH
• methylnitrosomočovina toxicita   MeSH
• monitorování životního prostředí MeSH
• mutageny toxicita   MeSH
• poškození DNA * MeSH
• tabák účinky léků   genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, U.S. Gov't, Non-P.H.S. MeSH
• srovnávací studie MeSH
• Názvy látek
• alkylační látky MeSH
• DNA rostlinná MeSH
• ethylmethansulfonát MeSH
• ethylnitrosomočovina MeSH
• methylmethansulfonát MeSH
• methylnitrosomočovina MeSH
• mutageny MeSH

A procedure for employing cultured tobacco cells (line TX1) in the SCGE assay was developed. The effect on DNA migration was studied in control and EMS-treated cells at different stages in their growth curve. The experimental parameters of treatment time and the unwinding time were analyzed in TX1 cells. With EMS in a concentration range from 0 to 30 mM the average median (+/-S.E.) tail moments ranged from 2.71+/-0.24 microm for the negative controls and increased in a direct concentration dependent manner to 57.89+/-4.13 for cells treated with 30 mM EMS. Nuclei isolated from TX1 cells and treated with EMS had a similar sensitivity as TX1 cells after EMS treatment. The plant cells express similar concentration-response curves for EMS as reported with mammalian (CHO) cells. This plant cell SCGE assay may prove to be a useful tool for the study of agricultural chemicals in specific plant cell types, to compare the response of mutagens in plant and animal cells and for basic research in genetic toxicology and DNA repair in plants.

• MeSH
• buněčné dělení účinky léků   genetika   MeSH
• buněčné jádro účinky léků   genetika   MeSH
• časové faktory MeSH
• elektroforéza metody   MeSH
• ethylmethansulfonát toxicita   MeSH
• jedovaté rostliny * MeSH
• kultivované buňky MeSH
• mutageny MeSH
• poškození DNA účinky léků   MeSH
• tabák cytologie   účinky léků   genetika   MeSH
• testy genotoxicity metody   MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, U.S. Gov't, Non-P.H.S. MeSH
• Názvy látek
• ethylmethansulfonát MeSH
• mutageny MeSH

The medium recovered from the tobacco cell suspension cultures (TX1MX) activated the promutagenic aromatic amine m-phenylenediamine (mPDA) and a macromolecular complex (gel) responsible for the arylamine activation was isolated from the medium. The gel formation and the role of the gel components in the plant activation of mPDA to products mutagenic in S. typhimurium YG1024 were studied. The activation of mPDA was caused by the peroxidases present in TX1MX. We demonstrated an association of the peroxidase activity and gel pectins. Formation of a stable mutagenic association of mPDA with the macromolecular material was observed. The data indicate that the gel isolated from TX1MX is the macromolecular component of the arylamine conjugate proposed in earlier work.

• MeSH
• biotransformace MeSH
• buněčné dělení MeSH
• buněčné kultury MeSH
• extracelulární matrix metabolismus   MeSH
• fenylendiaminy metabolismus   MeSH
• gely MeSH
• jedovaté rostliny * MeSH
• kultivační média speciální MeSH
• kyseliny uronové metabolismus   MeSH
• mutageny metabolismus   MeSH
• pektiny metabolismus   MeSH
• peroxidasy metabolismus   MeSH
• rostlinné proteiny biosyntéza   MeSH
• Salmonella typhimurium účinky léků   MeSH
• tabák enzymologie   metabolismus   MeSH
• testy genotoxicity MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, U.S. Gov't, Non-P.H.S. MeSH
• Názvy látek
• 3-phenylenediamine MeSH Prohlížeč
• fenylendiaminy MeSH
• gely MeSH
• kultivační média speciální MeSH
• kyseliny uronové MeSH
• mutageny MeSH
• pektiny MeSH
• peroxidasy MeSH
• rostlinné proteiny MeSH

Both tobacco cells in suspension and the medium recovered from the suspension cultures (TX1MX) activated the aromatic amine m-phenylenediamine (m-PDA) into a product that was mutagenic in Salmonella typhimurium TA98 and YG1024. Medium recovered from stationary-phase tobacco cell cultures exhibited the highest level of m-PDA activation. No cytochrome P-450 was detected in the activating medium. A high molecular weight matrix having the highest m-PDA activating capacity and associated with a substantial fraction of the total peroxidase activity was isolated by Centricon-100 ultrafiltration of TX1MX. The data suggest that the peroxidases present in the recovered cell culture medium or in the high molecular weight matrix are responsible for the plant activation of m-PDA.

• MeSH
• biotransformace MeSH
• fenylendiaminy metabolismus   MeSH
• jedovaté rostliny MeSH
• kultivační média MeSH
• kultivované buňky MeSH
• mutageny metabolismus   MeSH
• Salmonella typhimurium účinky léků   MeSH
• suspenze MeSH
• tabák metabolismus   MeSH
• ultrafiltrace MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, U.S. Gov't, Non-P.H.S. MeSH
• Názvy látek
• 3-phenylenediamine MeSH Prohlížeč
• fenylendiaminy MeSH
• kultivační média MeSH
• mutageny MeSH
• suspenze MeSH