Mechanism of comutagenesis of sodium arsenite withn-methyl-n-nitrosourea

Arsenic compounds are known carcinogens. Although many carcinogens are also mutagens, we have previously shown that sodium arsenite is not mutagenic at either the Na⁺/K⁺ ATPase orhprt locus in Chinese hamster V79 cells. It can, however, enhance UV-mutagenesis. We now confirm the nonmutagenicity of sodium arsenite in line G12, a pSV2gpt-transformed V79 (hprt ⁻) cell line, which is able to detect multilocus deletions in addition to point mutations and small deletions. The lack of arsenic mutagenicity has led to studies emphasizing its comutagenicity. Sodium arsenite at relatively nontoxic concentrations (5 μM for 24 h or 10 μM for 3 h) is comutagenic withN-methyl-N-nitrosourea (MMU) at thehprt locus in V79 cells. Using a nick translation assay, which measures DNA strand breaks by incorporating radioactive deoxyribonucleoside monophosphate at their 3′OH ends in permeabilized cells, we found that much more incorporation was seen in cells treated with MNU (4 mM, 15 min) followed by 3-h incubation with 10 μM sodium arsenite compared with cells exposed to the same MNU treatment followed by 3-h incubation without sodium arsenite. This result shows that in the presence of arsenite, strand breaks resulting from MNU or its repair accumulate over a 3-h period. We suggest that the repair of MNU-induced DNA lesions may be inhibited by arsenite either by affecting the incorporation of dNMPs into the MNU-damaged DNA template or by interfering with the ligation step.     

Performance of 133 compounds in the lambda prophage induction endpoint of the Microscreen assay and a comparison with S. typhimurium mutagenicity and rodent carcinogenicity assays.

The Microscreen assay was developed as a means of testing very small samples, as in complex mixture fractionation. It is a multi-endpoint assay which utilizes E. coli WP2s(lambda). Exposure takes place to serial dilutions of the test compound in microtitre wells (250 microliters) followed by sampling from wells in which growth has occurred ('non-toxic wells'). Although a number of different endpoints can be measured, only the prophage induction endpoint (the first one developed) has been extensively tested. Results with 133 compounds are presented. These include 111 compounds which have been tested in the S. typhimurium assay and 66 compounds for which both rodent bioassay and S. typhimurium assay data exists. The concordance for the Microscreen assay and the S. typhimurium assay was 71%. For this group of compounds, the sensitivity of the Microscreen assay in detecting carcinogens was 76% compared with 58% for the S. typhimurium assay. However, the S. typhimurium assay was somewhat more specific (69%) compared with the Microscreen (56%). The overall association between carcinogenicity and Microscreen results was statistically significant (p = 0.029), whereas for the S. typhimurium assay the association with carcinogenicity was non-significant (p = 0.086). The Microscreen assay was able to detect halogenated compounds better than the S. typhimurium assay. The Microscreen assay should prove useful in complex mixture fractionation, or in other situations where sample size is limiting.     

Differential interleukin 1 elaboration by unfractionated and density fractionated human alveolar macrophages and blood monocytes: relationship to cell maturity

The elaboration of interleukin 1 (IL 1) by mononuclear phagocytes is important in the regulation of human inflammatory and fibrotic reactions. Mononuclear phagocytes are morphologically and functionally heterogeneous cells. To further understand the processes controlling inflammation and fibrosis, in particular that in the human lung, we studied the elaboration of IL 1 by unfractionated and density-fractionated human alveolar macrophages and blood monocytes. Stimulated blood monocytes elaborated more IL 1 than stimulated alveolar macrophages. In addition, denser alveolar macrophages and blood monocytes elaborated more IL 1 than less dense alveolar macrophages and monocytes. Lastly, as monocytes matured in vitro, they lost their ability to elaborate IL 1 and became less dense. Thus, there is variability between and within mononuclear phagocyte cell populations in their ability to elaborate IL 1. These differences may result in part from differences in cell maturation.     

Virion orientation in cubic crystals of the human common cold virus HRV14

A new cubic crystal form (a = 445.1 A) of space group P23 is reported for human rhinovirus R14. There are four particles per unit cell, each situated on a crystallographic 3-fold axis. The orientation of these particles has been determined with a rotation function and their approximate positions have been derived from a Patterson map. The crystals diffract to at least 2.8 A resolution. Limitations to the possible surface features of the virus are set by a comparison of the cubic and orthorhombic crystal forms.     

The effects of bisulfite on growth and macromolecular synthesis in Escherichia coli

Bisulfite reversibly inhibits the growth of a variety of microorganisms and has been used as a preservative in foods and beverages for that reason. We have now measured macromolecule synthesis in Escherichia coli K12 after bisulfite treatment. RNA synthesis, the synthesis of total protein, and of an inducible enzyme, beta-galactosidase, stopped almost immediately upon addition of 2 mM (or higher concentrations) of bisulfite. These functions resumed after a lag whose duration depended on the concentration of bisulfite added. The synthesis of DNA was slowed upon bisulfite addition, but did not stop entirely. The inhibition of RNA synthesis by bisulfite took place in both stringent and relaxed strains of E. coli and was not relieved upon addition of chloramphenicol. Stringent control was therefore not involved in this effect. No effect on protein synthesis was observed in the cell-free system of E. coli (using poly(U) or MS2 RNA as messenger) at bisulfite concentrations up to 10 mM. Protein synthesis inhibition in vivo was apparently not due to a reaction of bisulfite with a component of this system. In additional experiments, RNA polymerase was not impaired by bisulfite, and the growth inhibition effect was shown to proceed in the presence of inhibitors of free radical chain reactions.     

Comutagenesis of sodium arsenite with ultraviolet radiation in Chinese hamster V79 cells

Summary Solar ultraviolet radiation has been associated with the induction of skin cancer. Recent studies have indicated that near-ultraviolet, especially UVB, is mutagenic. Exposure to trivalent inorganic arsenic compounds has also been associated with increased skin cancer prevalence. Trivalent arsenic compounds are not mutagenicper se, but are comutagenic with a number of cancer agents. Here, we test the hypothesis that arsenite enhances skin cancer via its comutagenic action with solar ultraviolet radiation. Irradiation of Chinese hamster V79 cells with UVA (360 nm), UVB (310 nm) and UVC (254 nm) caused a fluence-dependent increase in mutations at thehprt locus. On an energy basis, UVC was the most mutagenic and UVA the least. However, when expressed as a function of toxicity, UVB was more mutagenic than UVC. Nontoxic concentrations of arsenite increased the toxicity of UVA, UVB and UVC. Arsenite acted as a comutagen at the three wavelengths; however, higher concentrations of arsenite were required to produce a significant (P < 0.05) comutagenic response with UVB. The increased mutagenicity of UVB and UVA by arsenite may play a role in arsenite-related skin cancers.     

Functional anion binding sites in dogfish M4 lactate dehydrogenase

X-ray diffraction data have been collected from dogfish M₄ lactate dehydrogenase crystals in which ammonium sulfate had been exchanged by citrate at pH 6.0 and 7.8. Data were also collected from crystals which had been soaked in 0.1 m oxamate, a lactate dehydrogenase inhibitor. The difference electron density maps obtained have been interpreted in terms of two exchangeable anion binding sites, one at the active center and one between two subunits. The active center site is coincident with the substrate binding site in a ternary complex, while the subunit boundary site, which has been observed in several different forms of the enzyme, may be involved in stabilizing the tetramer.     

Pyrido[2,3-d]pyrimidines: Discovery and preliminary SAR of a novel series of DYRK1B and DYRK1A inhibitors

DYRK1B is a kinase over-expressed in certain cancer cells (including colon, ovarian, pancreatic, etc.). Recent publications have demonstrated inhibition of DYRK1B could be an attractive target for cancer therapy. From a data-mining effort, the team has discovered analogues of pyrido[2,3-d]pyrimidines as potent enantio-selective inhibitors of DYRK1B. Cells treated with a tool compound from this series showed the same cellular effects as down regulation of DYRK1B with siRNA. Such effects are consistent with the proposed mechanism of action. Progress of the SAR study is presented.     

Four putative entomopathogenic fungi of armoured scale insects on <Emphasis Type="Italic">Citrus</Emphasis> in Australia

This study led to the discovery of four putative entomopathogenic fungi of armoured scale insects on citrus trees in coastal New South Wales. Two of these species belong in Podonectria as P. coccicola (Ellis & Everh.) Petch (syn. Tetracrium coccicola (Höhn.) Ellis & Everh.) and P. novae-zelandiae Dingley. Members of this genus are grown in culture for the first time. Formerly placed in the Pleosporales, Tubeufiaceae, or more recently in the Tubeufiales, these species are herein placed in the new family Podonectriaceae fam. nov., Pleosporales. Another species is placed in the Hypocreales, Bionectriaceae as Clonostachys coccicola (J.A. Stev.) H.T. Dao comb. nov. (basionym Tubercularia coccicola J.A. Stev., syn. Nectria tuberculariae Petch). The fourth species is Myriangium citri Henn. (Myriangiales, Myriangiaceae). Each fungal species is characterized and the phylogenetic placement confirmed by molecular analyses of the ITS and 28 s rDNA regions. In addition, their biology is noted, including location of the fungi within tree canopies.