PubMed Health⌕ Search

Biomedical subjects

S L Barton

Publications and source records attributed to S L Barton.

13 recordsLinked to original sources
PubMed2001

Marker structure and recombination substrate environment influence conversion preference of broken and unbroken alleles in Saccharomyces cerevisiae.

Double-strand break (DSB)-induced gene conversion was investigated using plasmid x chromosome (P x C) and chromosomal direct-repeat recombination substrates with markers arranged such that functional (selected) products could not arise by longpatch mismatch repair initiated from the DSB. As seen previously with analogous substrates, these substrates yield products with discontinuous conversion tracts, albeit at low frequency. Most conversion tracts were of minimum length, suggesting that heteroduplex DNA (hDNA) is limiting, or that co-repair imposes selective pressure against products with more extensive hDNA. When functional products can arise by long-patch mismatch repair, the broken allele is converted in nearly all products. In contrast, in the absence of long-patch mismatch repair, unbroken alleles are frequently converted, and we show that such conversion depends on both marker structure (i.e., long palindromic vs. nonpalindromic insertions) and the chromosomal environment of the recombination substrate. We propose that conversion of unbroken alleles is largely a consequence of the segregation of unrepaired markers, and that differences in mismatch repair efficiency underlie the observed effects of marker structure and chromosome environment on allele conversion preference.

Alleles↗
PubMed2001

Homologous recombinational repair of double-strand breaks in yeast is enhanced by MAT heterozygosity through yKU-dependent and -independent mechanisms.

DNA double-strand breaks (DSBs) are repaired by homologous recombination (HR) and nonhomologous end-joining (NHEJ). NHEJ in yeast chromosomes has been observed only when HR is blocked, as in rad52 mutants or in the absence of a homologous repair template. We detected yKu70p-dependent imprecise NHEJ at a frequency of approximately 0.1% in HR-competent Rad+ haploid cells. Interestingly, yku70 mutation increased DSB-induced HR between direct repeats by 1.3-fold in a haploid strain and by 1.5-fold in a MAT homozygous (a/a) diploid, but yku70 had no effect on HR in a MAT heterozygous (a/alpha) diploid. yku70 might increase HR because it eliminates the competing precise NHEJ (religation) pathway and/or because yKu70p interferes directly or indirectly with HR. Despite the yku70-dependent increase in a/a cells, HR remained 2-fold lower than in a/alpha cells. Cell survival was also lower in a/a cells and correlated with the reduction in HR. These results indicate that MAT heterozygosity enhances DSB-induced HR by yKu-dependent and -independent mechanisms, with the latter mechanism promoting cell survival. Surprisingly, yku70 strains survived a DSB slightly better than wild type. We propose that this reflects enhanced HR, not by elimination of precise NHEJ since this pathway produces viable products, but by elimination of yKu-dependent interference of HR.

Alleles↗
PubMed1997

Carcinogenic polycyclic aromatic hydrocarbons increase intracellular Ca2+ and cell proliferation in primary human mammary epithelial cells.

Previous studies have shown that polycyclic aromatic hydrocarbons (PAHs) mobilize intracellular Ca2+ in human T cells by inositol trisphosphate-dependent mechanisms resulting from activation of phospholipase C-gamma by SRC-related protein tyrosine kinases, thereby mimicking antigen-receptor activation. Ca2+ appears to play an important second messenger role in growth factor control of cell proliferation in human mammary epithelial cells (HMEC), such as the epidermal growth factor receptor pathway. The purpose of the present studies was to determine if PAHs are able to increase intracellular Ca2+ in primary cultures of HMEC and increase cell proliferation. Two carcinogenic and two non-carcinogenic PAHs were tested for their ability to increase intracellular Ca2+ in HMEC. The carcinogenic PAHs dimethylbenz[a]anthracene (DMBA) and benzo[a] pyrene (BaP) were able to cause Ca2+ elevation in HMEC at early time points (2 h) and caused sustained alterations in Ca2+ homeostasis (18 h). DMBA showed maximal effects at early time points (2 h), while BaP showed maximal effects on sustained Ca2+ (18 h). 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD), a potent dioxin and tumor promoter, produced maximal Ca2+ elevation at 2 h, with a return to near baseline levels by 6 h. The non-carcinogenic PAHs benzo[e]pyrene and anthracene did not significantly alter intracellular Ca2+ at any time point. alpha-Naphthoflavone significantly reduced the Ca2+ response induced by BaP treatment, but not by DMBA or TCDD, suggesting that P450 1A or 1B metabolism of BaP may be important in the sustained Ca2+ elevating response. In evaluating the effects of BaP on HMEC proliferation, BaP was found to increase the number of cells recovered after 4 days in culture in the absence or presence of various concentrations of epidermal growth factor. These studies provide initial evidence that Ca2+ signaling may be associated with mitogenesis in HMEC, which may play a role in tumor promotion and progression produced by PAHs.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1996

GM-CSF secretion in primary cultures of normal and cancerous human renal cells.

Rates of secretion of granulocyte-macrophage colony stimulating factor (GM-CSF) were measured in 50 primary cell cultures derived from cancerous and normal human kidneys. Mean rates of GM-CSF secretion measured by TF-1 cell proliferation assay (N = 21) and by ELISA (N = 31) were 2.5 and 7.8 ng/10(6) cells/24 hr, respectively. There was no significant difference between the mean rates of GM-CSF secretion by cancerous and normal renal cells. GM-CSF was also secreted by primary renal cell cultures grown in serum-free medium and by renal cell lines. GM-CSF mRNA was detected by RT-PCR in cultured renal cells, but not in undissociated kidney tissue. Rates of GM-CSF secretion were reduced up to 99% under conditions where the cellular density or substratum more closely resembled the in vivo environment. Some cultured human renal carcinoma cells (RCC) secreted GM-CSF at levels that occasionally overlapped the levels produced by the GM-CSF gene-modified human RCC vaccine now in phase I trial. The data indicate that GM-CSF is not expressed in vivo, and that stable GM-CSF secretion is induced by the dissociation and culture of human renal cells.

Adenocarcinoma↗
PubMed1993

DMBA induces programmed cell death (apoptosis) in the A20.1 murine B cell lymphoma.

The mechanism by which 7,12-dimethylbenz[a]anthracene (DMBA) produces cytotoxicity in lymphocytes was investigated in these studies using the murine A20.1 B cell lymphoma. Results show that in vitro exposure of these cells to 10-30 microM DMBA for 4 hr produced an increase in intracellular Ca2+, DNA fragmentation, and subsequent cell death. Elevation of Ca2+ and DNA fragmentation induced by DMBA were greatly pronounced when the A20.1 cells were exposed at high cell density (10(7) cells/ml). DMBA-induced DNA fragmentation and cell death were inhibited by coexposure of A20.1 cells to a calcium chelator (EDTA), a general nuclease and polymerase inhibitor (aurintricarboxylic acid), and a protein synthesis inhibitor (cycloheximide). These agents have been previously shown to inhibit apoptosis in lymphocytes and other cells exposed to chemical agents. We also found that cyclosporin A, an inhibitor of Ca(2+)-dependent pathways of T and B cell activation, prevented apoptosis in the A20.1 cell line. These results demonstrate that DMBA induces programmed cell death (apoptosis) in the A20.1 murine B cell lymphoma by Ca(2+)-dependent pathways. The increased sensitivity of A20.1 at high cell density to Ca2+ elevation and DNA fragmentation suggests that cell to cell interactions may also be important in this process.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1993

A histidine decarboxylase-like mRNA is involved in tomato fruit ripening.

DNA sequencing of a tomato ripening-related cDNA, TOM 92, revealed an open reading frame with homology to several pyridoxal 5'-phosphate histidine decarboxylases, containing the conserved amino acid residues known to bind pyridoxal phosphate and alpha-fluoromethylhistidine, an inhibitor of enzyme activity. TOM 92 mRNA accumulated during early fruit ripening and then declined. Fruit of the ripening-impaired tomato mutant, ripening inhibitor (rin), did not accumulate TOM 92 mRNA, and its accumulation was not restored by treatment of fruit with ethylene. The TOM 92 mRNA was not detected in tomato leaves and unripe fruit.

Amino Acid Sequence↗
PubMed1991

Suppression of local gut-associated and splenic mitogen responsiveness of lymphoid cells following oral exposure of B6C3F1 mice to 7,12-dimethylbenz[a]anthracene.

In these studies, the mitogen responsiveness of lymphocytes obtained from local gut-associated lymphoid tissues (GALT) and the spleen were evaluated following a 5-day exposure to 7,12-dimethylbenz(a)anthracene (DMBA) at doses of 50 or 150 mg/kg. Phytohemagglutinin and lipopolysaccharide (LPS) responses were suppressed in all lymphoid tissues studied. However, the LPS response in mesenteric lymph nodes and Peyer's Patches seemed to be the most sensitive indicator of immunotoxicity, indicating that B cells appear to be particularly sensitive to DMBA toxicity in the GALT. These studies demonstrate that both splenic tissues and GALT are important targets of immunotoxicity following oral administration of DMBA. Based upon these and past studies we conclude that the total administered dose of DMBA is a more important determinant of immunotoxicity than the length of exposure.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1990

Inhibition of lymphocyte activation in splenic and gut-associated lymphoid tissues following oral exposure of mice to 7,12-dimethylbenz[a]anthracene.

The hypothesis that 7,12-dimethyl-benz[a]anthracene (DMBA) suppresses immune function in mice via an inhibition of lymphocyte activation was examined in these studies. Daily exposure of B6C3F1 mice to DMBA (cumulative doses of 1.4 to 140 mg/kg) via the oral route for 14 days was found to inhibit phytohemagglutinin (PHA) and lipopolysaccharide mitogen responses in lymphoid cells obtained from the spleen. Peyer's Patches, and mesenteric lymph nodes. The 14 mg/kg cumulative dose of DMBA produced no significant decrease in the number of recovered viable cells, yet mitogen responses were suppressed by approximately 50% in the spleen and mesenteric lymph nodes, and by greater than 70% in the Peyer's Patches. DMBA inhibited PHA-induced Ca+2 mobilization measured by flow cytometry in each of these three lymphoid tissues. There was no change in the percentage of T cells recovered from the spleen, mesenteric lymph nodes, or Peyer's Patches. Peyer's Patch lymphocytes obtained from the GI tract appeared to be slightly more sensitive to inhibition of mitogen responsiveness and perhaps Ca+2 mobilization, potentially due to the oral route of exposure to DMBA. These studies provide evidence that DMBA inhibits early events associated with lymphocyte activation in mice.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1988

Persistent suppression of humoral immunity produced by 7,12-dimethylbenz(A)anthracene (DMBA) in B6C3F1 mice: correlation with changes in spleen cell surface markers detected by flow cytometry.

The purpose of these studies was to examine the effects of DMBA on subpopulations of splenocytes obtained from B6C3F1 mice, using cell surface markers defined by monoclonal antibodies and multiparameter flow cytometry. Changes were correlated with alterations in humoral immune function assessed in vitro. Mice were treated for 10 days during a 2 week period by subcutaneous (s.c.) injections of DMBA in corn oil at doses of 0.5, 5 and 10 micrograms/g/day (5-100 micrograms/g total dose). Four mice from each exposure group and an additional corn oil control group of mice were studied at 4 and 8 weeks following the last injection with DMBA. These studies demonstrated a dose-dependent decrease in the total number and percentage of spleen cells expressing B-cell markers (mu heavy chain, kappa light chain and 14.8 antigen) as well as T-cell markers (Thy 1.2, Lyt-1 and Lyt-2). The percentage of splenocytes expressing Mac-1 was increased by DMBA. Helper T-cells appeared to be a very sensitive population of spleen cells to DMBA exposure, as suggested by a decrease in the number and percentage of Lyt-1 positive cells recovered from the spleen 4 weeks after exposure to DMBA. DMBA produced a dose-dependent suppression of the in vitro primary humoral immune responses to SRBC, TNP-Ficoll and TNP-LPS. The fact that a functional suppression of humoral immunity was accompanied by a decrease in the number of mature B-cells and T-cells in the spleen suggests that cell surface markers may be useful indicators of immunotoxicity in animals receiving DMBA in sub-chronic studies.

9,10-Dimethyl-1,2-benzanthracene↗