Use of UV endonuclease from Micrococcus luteus to monitor the progress of DNA repair in UV-irradiated human cells

A sensitive enzymatic assay has been developed to follow the progress of NDA repair in human cells exposed to UV radiation. The assay employs an endonuclease selectively active at sites containing pyrimidine dimers in UV-damaged DNA. Primary fibroblasts are exposed to 254 nm radiation and incubated for specified times, their radioactivity labelled DNA is isolated and treated with a UV endonuclease extensively purified from Micrococcus luteus. Endonuclease-susceptible site remaining in the DNA are subsequently observed as single-strand scissions by sedimentation in alkaline sucrose gradients. In comparison to the situation with excision-proficient normal cells, those derived from patients suffering from either the classical or the De Sanctis-Cacchione clinical form of Xeroderma pigmentosum (XP) exhibit a marked diminution in the rate of disappearance of nuclease-susceptible lesions with time of post-UV incubation.     

Posttranslational Modifications in Connexins and Pannexins

Posttranslational modification is a common cellular process that is used by cells to ensure a particular protein function. This can happen in a variety of ways, e.g., from the addition of phosphates or sugar residues to a particular amino acid, ensuring proper protein life cycle and function. In this review, we assess the evidence for ubiquitination, glycosylation, phosphorylation, S-nitrosylation as well as other modifications in connexins and pannexin proteins. Based on the literature, we find that posttranslational modifications are an important component of connexin and pannexin regulation.     

Novel, potent, selective, and orally bioavailable human betaII-tryptase inhibitors

The synthesis of novel [1,2,4]oxadiazoles and their structure-activity relationship (SAR) for the inhibition of tryptase and related serine proteases is presented. Elaboration of the P'-side afforded potent, selective, and orally bioavailable tryptase inhibitors.     

Cellular and molecular analysis of plasmodium development in Physarum.

The development of an amoeba into a plasmodium involves extensive changes in cellular organisation and gene expression. The genetic basis of a number of recessive mutations that block plasmodium development has been elucidated. The stage at which development becomes abnormal has been determined for all the mutants, as has the terminal phenotype. In order to investigate the changes in gene expression that accompany plasmodium development, a cDNA library has been made using RNA isolated from cell populations in which development was occurring.     

PHYSIOLOGICAL INDICATORS OF NUTRIENT DEFICIENCY IN CLADOPHORA (CHLOROPHYTA) IN THE CLARK FORK OF THE COLUMBIA RIVER,MONTANA1

Cellular nutrient concentrations and nutrient uptake rates of Cladophora glomerata (L.) Kuetzing were determined during summer and fall in 1989–1990 at a site on the upper Clark Fork of the Columbia River, Montana. Both physiological tests indicated that Cladophora growth is likely to be limited by nitrogen during late summer-early fall. Maximum uptake rates of ammonia-N and nitrate-N were 5935–6991 and 507–984 μg · g DW^?1· h^?1, respectively, during July–October when dissolved inorganic nitrogen (DIN) concentrations in the river were less than 10 μg · L^?1. During November-December, when DIN was 72–376 μg · L^?1, maximum ammonia-N uptake was 1137–1633 μg · g DW^?1· h^?1 and maximum nitrate-N uptake was 0–196 μg · g DW^?1· h^?1. Cellular nitrogen during summer–early fall was 0.78–1.80% of Cladophora dry weight, frequently at or below 1.1%, a level suggested as a critical minimum N concentration for maximum growth. In contrast, cellular P was 0.18–0.36% of dry weight, 3–6 times the suggested critical P concentration of 0.06%. Molar ratios of cellular N:P (< 16:1) and DIN: SRP (< 4:1) during late summer-early fall also indicated potential N limitation. Cellular N and P from Cladophora collected from a second site influenced by a municipal wastewater discharge in 1990 displayed similar seasonal trends. At both sites, seasonal fluctuations in DIN were closely tracked by changes in cellular N, Cellular P, however, increased through the growing season despite declining levels of SRP in the river.     

Influence of treatment temperature on the genotoxic effects of cisplatin in CHO cells: cytotoxicity, mutagenicity and induction of lesions in DNA

In cells exposed in vitro to the cytotoxic and mutagenic antitumor drug cisplatin (cis-Pt(NH3)2Cl2), various adducts with nuclear DNA are formed. A comparative study was made of the influence of temperature variation during treatment of cultured Chinese hamster ovary (CHO) cells with cisplatin on cytotoxicity, mutation induction and Pt-DNA adduct formation. Before and after treatment (1 h at 32, 37 or 40 degrees C) cells were kept at 37 degrees C. Cytotoxicity increased with temperature; D0 values were 29.6 +/- 1.6, 21.1 +/- 1.2 and 11.4 +/- 0.6 microM at 32, 37 and 40 degrees C, respectively. Pt-DNA binding to DNA at 40 degrees C was 2.0 (+/- 0.3) times as high as at 32 degrees C. This factor remained practically constant over a 24-h post-treatment incubation of the cells, during which about 60% of DNA-bound Pt were removed. As the increase in cytotoxicity between 32 and 40 degrees C was roughly in proportion to that in Pt binding, no substantial changes in the spectrum of adducts appeared to occur. The induction of DNA interstrand cross-links, studied at 32 and 40 degrees C, varied linearly with dose. Influence of temperature on cross-link formation was comparable to that on total Pt binding. Amounts of cross-links highly increased during 24 h after treatment. Plots of cross-links against survival after treatments at 32 and 40 degrees C almost coincided. Induction of 6-thioguanine-resistant (HGPRT) mutants at various cisplatin concentrations did not show a clear temperature dependency. Consequently, equitoxic treatments were significantly more mutagenic at 32 degrees C than at 40 degrees C, the opposite of what has been reported for E. coli.