Lipid peroxidation and the polymerase activities of liver chromatin fractions in rats during aging

Lipids, which enter the composition of actively transcribed and repressed chromatin fractions are found to undergo a peroxidation. This process decreased with aging and more pronounced in actively transcribed chromatin fraction. A decreased activity of DNA polymerase beta and increased activity of RNA polymerase I in this chromatin fraction with aging were observed. It is assumed that observed changes of genome function of old animals may be caused by decreased peroxidation of chromatin lipids.     

Purification and properties of tyrosinases from Vibrio tyrosinaticus

Rat liver chromatin which has been briefly sonicated is fractionated by treatment with low concentrations of magnesium ion. At 1.5 mm Mg²⁺, where approximately 20–25% of the chromatin remains soluble after low-speed centrifugation, chemical and physical analysis of the Mg-soluble and Mg-insoluble chromatin fractions show that the fractions possess markedly different properties. The Mg-soluble chromatin has more protein and RNA than the Mg-insoluble chromatin. The histone composition of the two fractions as shown by electrophoretic analysis is similar, but many of the acidic proteins are qualitatively and quantitatively different. The molecular weight of the Mg-soluble chromatin is less than that of the insoluble chromatin based on sedimentation behavior and gel filtration experiments. The soluble chromatin has nearly twice the template activity for RNA synthesis in vitro with added RNA polymerase as the Mg-insoluble chromatin and contains approximately 80% of the in vivo rapidly labeled RNA found in the total chromatin preparation. In addition the Mg-soluble chromatin has a significantly greater amount of “accessible” DNA (62%) as measured by polylysine binding than Mg-insoluble chromatin (48%). The data suggest that (a) fractionation of chromatin preparations can be achieved by titration with Mg²⁺, and (b) chromatin soluble in low concentrations of Mg²⁺ may be enriched in actively transcribed portions of the genome.     

Epigenetic regulation of transcription by RNA polymerase III

Chromatin participates actively in all DNA transactions and all phenomena directly under the influence of chromatin are explained by epigenetic mechanisms. The genes transcribed by RNA polymerase (pol) III are generally found in regions free of nucleosomes, the structural units of chromatin. Yet, histone modifications and positions of nucleosomes in the gene flanking regions have been reported to show direct correlation with activity status of these genes. Gene-specific as well as genome-wide studies have also revealed association of several epigenetic components with pol III-transcribed genes. This review presents a summary of the research in past many years, which have gathered enough evidence to conclude that pol III-transcribed genes are important components of an epigenome.     

The endogenous nuclease sensitivity of repaired DNA in human fibroblasts

The limited DNA excision repair that occurs in the chromatin of UV-irradiated growth arrested cells isolated from a xeroderma pigmentosum (XP) complementation group C patient is clustered in localized regions. The repaired DNA was found to be more sensitive to nicking by endogenous nucleases than the bulk of the DNA. The extra-sensitivity does not change with increasing amounts of DNA damage or repair activity in the locally-repaired regions and is retained through a 24-h chase period. We suggest that these results are due to the occurrence of DNA repair limited to pre-existing, non-transient chromatin fractions that contain actively transcribed DNA. A similar extra-sensitivity of repaired DNA was not detected in cells of normal or XP complementation group A strains that exhibit either normal or limited repair located randomly throughout their genomes. The association between endogenous nuclease sensitivity and clustered repair probably defines a normal excision repair pathway that is specific for selected chromatin domains. The repair defect in XP-C strains may be one in pathways targeted for other endogenous nuclease-resistant domains.     

Arachidonic acid hydroperoxide stimulates lipid peroxidation in rat liver nuclei and chromatin fractions

Arachidonic acid, the most abundant polyunsaturated fatty acid in rat liver nuclei phospholipids is a major target of free radical attack, which induces lipid peroxidation. The non-enzymatic lipid peroxidation process in intact rat liver nuclei and in several chromatin fractions indicated that the most sensitive fatty acid for peroxidation is arachidonic acid C20:4 n-6. In this study, the effect of different amounts of arachidonic acid hydroperoxide on the lipid peroxidation of rat liver nuclei and chromatin fractions was studied; rat liver nuclei and chromatin fractions deprived of exogenous added hydroperoxide were utilized as control. The addition of arachidonic acid hydroperoxide to liver nuclei produces a marked increase in light emission that was hydroperoxide concentration dependent. The maximal peak of chemiluminescence displayed by the different chromatin fractions analyzed was observed between 20 and 80 min of incubation. The highest value of light emission was displayed by the high-density chromatin fractions, the 27.5 K fraction showed intermediate values of light emission, whereas the lowest density fraction produced very low chemiluminescence. A high correlation between arachidonic acid hydroperoxide concentration and chemiluminescence in the different chromatin fractions was observed. AC is Members of Carrera del Investigador Científico, Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Argentina.