Metabolism of the mutagen MeIQx in vivo: metabolite screening by liquid chromatography-thermospray mass spectrometry

2-Amino-3,8-dimethylimidazo(4,5-f)quinoxaline (MeIQx) is a potent mutagen found in cooked food. MeIQx and its isotopically labelled (13C, 15N2 and 14C) analogues were synthesised and used for metabolic studies in vivo. An equimolar mixture of MeIQx and its 13C, 15N2 stable isotope labelled analogue (containing tracer amounts of 14C-MeIQx) was given intraperitoneally to mice. Some 67% of the radioactivity was eliminated in urine and faeces within 24h. Four radiolabelled species were observed when urine was analysed by HPLC, corresponding to unchanged MeIQx and three more polar metabolites. Urine was analysed directly by HPLC-thermospray mass spectrometry. Four signals were observed containing the characteristic 1:1 isotopic doublet, corresponding to unchanged MeIQx, an MeIQx glucuronide, and two uncharacterized metabolites.     

Chinese hamster metaphase chromosomes isolated under physiological conditions. A partial characterization of associated non-histone proteins and protein cores

In a previous report [2] we have described a non-histone protein core which could be isolated from Chinese hamster metaphase chromosomes. This core structure maintained the overall morphology of the metaphase chromosome even after removal of all of the histones, together with many of the non-histone proteins and the bulk of the DNA. As part of our work on the characterization of these core structures, we have developed a novel procedure for the isolation of metaphase chromosomes which avoids the use of high pH buffers and hexylene glycol, as well as eliminating the numerous centrifugation and resuspension steps previously employed. Chromosome cores prepared by 2 M NaCl extraction and DNase I digestion from metaphase chromosomes isolated under these more gentle, quasi-physiological conditions, are shown to contain a relatively simple subset of non-histone proteins. One-dimensional SDS-polyacrylamide gel electrophoresis shows two major groups of polypeptides having molecular weights 48 000-52 000 and 65 000-72 000 D respectively, with similarities in mobilities to the nuclear pore complex-lamina polypeptides and tubulins. However, more detailed analysis by two-dimensional gel electrophoresis and peptide mapping has failed to detect these proteins. A 52 000 D polypeptide component of the core is tentatively identified as the intermediate filament protein vimentin. The in vivo significance of chromosome cores is discussed.     

The isolation, characterization and partial sequence of a peptide rich in glutamic acid and aspartic acid (HGA-2 peptide) from calf thymus non-histone chromosomal protein HMG 2. Comparison with a similar peptide (HGA-1 peptide) from calf thymus non-histone chromosomal protein HMG 1.

A 41-residue peptide (HGA-2) containing a continuous sequence of 35 glutamic and aspartic residues was isolated from non-histone chromosomal protein HMG 2. This highly acidic peptide is compared with a similar peptide (HGA-1) isolated from non-histone chromosomal protein HMG 1.     

Phytoalexin resveratrol attenuates the mutagenicity of the heterocyclic amines 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine and 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline

Resveratrol is a phytoalexin, that belongs to a family of naturally occurring stilbenes. It has been reported that resveratrol can inhibit chemical carcinogenesis in experimental animals and although the mechanisms involved are unknown, an anti-mutagen mechanism has been proposed. We have explored this hypothesis using mutagenicity assays based on bacterial (Salmonella typhimurium) and eukaryotic cells (Chinese hamster V79 cells). We found resveratrol to be potent in both systems, blocking the mutagenicity of the food-derived heterocyclic amines (HA) 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) at micromolar concentrations. Furthermore, in cells capable of activating 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine to cytotoxic derivatives, resveratrol was able to attenuate cytotoxicity. Paradoxically, in cells lacking the ability to activate PhIP, resveratrol itself was toxic and co-incubation with PhIP reduced this toxicity. Our data confirm the potent anti-mutagenic activity of resveratrol and support its potential as a chemopreventative.     

Regulation of virulence and antibiotic resistance by two-component regulatory systems in Pseudomonas aeruginosa

The Gram-negative opportunistic pathogen Pseudomonas aeruginosa ubiquitously inhabits soil and water habitats and also causes serious, often antibiotic resistant, infections in immunocompromised patients (e.g. cystic fibrosis). This versatility is mediated in part by a large repertoire of two-component regulatory systems that appear instrumental in the regulation of both virulence processes and resistance to antimicrobials. Major two-component regulatory system proteins demonstrated to regulate these diverse processes include PhoP–PhoQ, GacA–GacS, RetS, LadS, and AlgR, among others. Here, we summarize the current body of knowledge of these and other two-component systems that provides insight into the complex regulation of virulence and resistance in P. aeruginosa .     

A simple method for the extraction and preservation of an undisturbed root system from a soil

Summary A simple method is described for the presentation in three dimensions of an undisturbed root system which is extracted from a soil core. A nylon mesh is threaded into the soil core in order to support the root system whilst the soil is removed by washing. The support for the root system provided by the nylon mesh is replaced by gelatin, the whole being transferred to a perspex box. In this way, an undisturbed root system may be studied, photographed, and preserved.     

Induction of xanthine oxidase and depression of cytochrome P-450 by interferon inducers: genetic difference in the responses of mice

Interferon and interferon inducing agents depress hepatic cytochrome P-450 systems. They also induce hepatic xanthine oxidase activity. It has been suggested that free radicals produced by xanthine oxidase may cause the loss of P-450. High titers of serum interferon are induced by poly IC (poly riboinosinic acid.polyribocytidylic acid) in both C57Bl/6J and C3H/HeJ mice; Newcastle disease virus (NDV) induces a high titer of interferon in C57Bl/6J mice but not in C3H/HeJ mice. The induction of xanthine oxidase activity by NDV in C3H/HeJ mice was less than half that seen in C57Bl/6J mice, thus demonstrating a relationship between the induction of xanthine oxidase, the depression of P-450 and a genetically determined difference in responsiveness of mice to interferon inducers.     

The effects of histones H1o,H5 and HMG proteins on cell division of cultured murine erythroleukemia cells

Summary In the present study the effect of histones H1^o and H5, and the nonhistone chromatin proteins HMG 1, 2, 14 and 17 (the high mobility group proteins), as well as the acidic peptide fragments of HMG 1 and 2 and polyglutamate, on cell division and differentation of cultured murine erythroleukemia (Friend) cells has been investigated. It was found that histones H1^o and H5, the acidic peptide fragments of HMG 1 and 2, HMG 14 and 17 and sodium polyglutamate stimulated cell division at a concentration of 10 g/ml. None of the H1^o, H5 or HMG protein preparations induced hemoglobin synthesis, as judged by benzidine staining.     

The isolation and partial sequence of peptides produced by cyanogen bromide cleavage of calf thymus non-histone chromosomal high-mobility-group protein 2. Sequence homology with non-histone chromosomal high-mobility-group protein 1.

Peptides produced by CNBr cleavage of non-histone chromosomal protein HMG 2 (CNBr peptides) were isolated and characterized, and their partial sequences were determined. The present sequence data account for over half of the sequence of the protein HMG (high-mobility-group) 2 molecule, and, together with previously published results, provide interesting information on the charge distribution within the molecule. Comparison of the CNBr-peptide-sequence data for protein HMG 2 with the previously published data on the CNBr peptides from protein HMG 1 reveals extensive sequence homology between the two proteins. Detailed evidence for the amino acid-sequence data has been deposited as Supplementary Publication SUP 50095 (6 pages) at the British Library Lending Division, Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies may be obtained on the terms given in Biochem. J. (1978) 169, 5.