Elena I. Georgieva

Alterations in p53, BRCA1, ATM, PIK3CA, and HER2 genes and their effect in modifying clinicopathological characteristics and overall survival of Bulgarian patients with breast cancer

PurposeThough p53, BRCA1, ATM, PIK3CA, and HER2 genes are shown to be involved in various aspects of breast carcinogenesis, their functional relationship and clinical value are still disputable. We investigated the genetic status or expression profile of these genes to further elucidate their clinical significance.MethodsPCR-SSCP-Sequencing of p53, BRCA1, ATM, and PIK3CA was performed in 145 Bulgarian patients with sporadic breast cancer. Expression profiles of HER2 were determined by ICH and CISH. Relationship between mutations and clinicopathological characteristics was evaluated by Chi-squared and Fisher’s exact tests. Multivariate Cox proportional hazard test and Kaplan–Meier analysis were used to evaluate differences in overall survival between groups.ResultsThe frequency of p53 (22.07%), BRCA1 (0.69%), ATM (7.59%), and PIK3CA (31.25%) alterations and HER2 (21.21%) overexpression was estimated. Mutated p53 was associated with tumor size (Pxa0=xa00.033) and grade of malignancy (Pxa0=xa00.001), ATM—with grade of malignancy (Pxa0=xa00.032), and PIK3CA—with PR-positive tumors (Pxa0=xa00.047). HER2 overexpression correlated with age of diagnosis (Pxa0=xa00.009), tumor size (Pxa0=xa00.0004), and ER expression (Pxa0=xa00.011). Univariate survival analysis showed that mutated p53 is an indicator for worse outcome (Pxa0=xa00.041). Combination of two genetic abnormalities did not correlate with more aggressive carcinogenesis and worse overall survival.ConclusionsOur data indicated that p53, BRCA1, ATM, PIK3CA, and HER2 alterations specifically correlate with clinicopathological characteristics of Bulgarian patients with breast cancer. Of these genes, only mutated p53 showed significant, though not independent, negative effect on overall survival.

Distribution of high mobility group and other acid-soluble proteins in fractionated chromatin

Chromatin was fractionated by digestion with deoxyribonuclease II and precipitation with MgCl2. The Mg2+-soluble fraction, known to be enriched in transcribed DNA sequences, was enriched also in high mobility group proteins 1 and 2 and contained almost all other acid-soluble nonhistone proteins.

A Short-range Gradient of Histone H3 Acetylation and Tup1p Redistribution at the Promoter of the Saccharomyces cerevisiae SUC2 Gene*

Chromatin immunoprecipitation assays are used to map H3 and H4 acetylation over the promoter nucleosomes and the coding region of the Saccharomyces cerevisiae SUC2 gene, under repressed and derepressed conditions, using wild type and mutant strains. In wild type cells, a high level of H3 acetylation at the distal end of the promoter drops sharply toward the proximal nucleosome that covers the TATA box, a gradient that become even steeper on derepression. In contrast, substantial H4 acetylation shows no such gradient and extends into the coding region. Overall levels of both H3 and H4 acetylation rise on derepression. Mutation of GCN5 or SNF2 lead to substantially reduced SUC2 expression; in gnc5 there is no reduction in basal H3 acetylation, but large reductions occur on derepression. SNF2 mutation has little effect on H3 acetylation, so SAGA and SWI/SNF recruitment seem to be independent events. H4 acetylation is little affected by either GCN5 or SNF2 mutation. In a double snf2/gcn5 mutant (very low SUC2 expression), H3 acetylation is at the minimal level, but H4 acetylation remains largely unaffected. Transcription is thus linked to H3 but not H4 acetylation. Chromatin immunoprecipitation assays show that Tup1p is evenly distributed over the four promoter nucleosomes in repressed wild type cells but redistributes upstream on derepression, a movement probably linked to its conversion from a repressor to an activator.

Role of superoxide dismutase in the oxidation of N-alkanes by yeasts

Yeast microorganisms from Candida genus are investigated for their superoxide dismutase (SOD) and catalase activity during cultivation on N-alkanes. The later caused a considerable increase of Cu/Zn SOD activity of yeast cells in comparison with glucose. A correlation between SOD and catalase activity existed. It is further observed that cells of Candida lipolytica 68-72 which contain a high level of Cu/Zn SOD were more resistant to lethality of exogenous O2-. An over-production of Cu/Zn SOD during the assimilation of N-alkanes by yeasts is also connected to their considerable resistance to increased concentrations of Cu2+ and Zn2+ ions in the nutrient medium. The results are consistent with the assumption that the enhanced resistance of yeast cells to O2- and high concentrations of Cu2+ and Zn(2+)-ions are due to the increased activity of Cu/Zn SOD and that SOD is involved in the protection of some cellular components. Polyacrylamide gel electrophoresis of Candida lipolytica cell-free extracts revealed the same chromatic bands of SOD activity under growth on glucose and N-alkanes. The type of the carbon source used from yeast cells as a single source of carbon and energy had no influence on the SOD profile of the cell.

Distribution of acetylated forms of nucleosomal histones in fractionated chromatin

Abstract Hyperacetylated chromatin was isolated from Ehrlich ascites tumor cells grown in n -butyrate containing medium and fractionated by mild digestion with deoxyribonuclease II and precipitation with MgCl 2 . The most highly acetylated forms of histones H4 and H3 as well as the acetylated subspecies of H2B were found almost entirely associated with the putatively active Mg 2+ -soluble chromatin fraction. The Mg 2+ -insoluble fraction contained mainly mono- and diacetylated molecules of H4 and H3.