Decline of nucleotide excision repair capacity in aging <Emphasis Type="Italic">Caenorhabditis elegans</Emphasis>

Background  

Caenorhabditis elegans is an important model for the study of DNA damage and repair related processes such as aging, neurodegeneration, and carcinogenesis. However, DNA repair is poorly characterized in this organism. We adapted a quantitative polymerase chain reaction assay to characterize repair of DNA damage induced by ultraviolet type C (UVC) radiation in C. elegans, and then tested whether DNA repair rates were affected by age in adults.     

Reductive debromination of the commercial polybrominated biphenyl mixture firemaster BP6 by anaerobic microorganisms from sediments.

Anaerobic microorganisms eluted from three sediments, one contaminated with polybrominated biphenyls (PBBs) and two contaminated with polychlorinated biphenyls, were compared for their ability to debrominate the commercial PBB mixture Firemaster. These microorganisms were incubated with reduced anaerobic mineral medium and noncontaminated sediment amended with Firemaster. Firemaster averages six bromines per biphenyl molecule; four of the bromines are substituted in the meta or para position. The inocula from all three sources were able to debrominate the meta and para positions. Microorganisms from the Pine River (St. Louis, Mich.) contaminated with Firemaster, the Hudson River (Hudson Falls, N.Y.) contaminated with Aroclor 1242, and Silver Lake (Pittsfield, Mass.) contaminated with Aroclor 1260 removed 32, 12, and 3% of the meta plus para bromines, respectively, after 32 weeks of incubation. This suggests that previous environmental exposure to PBBs enhances the debromination capability of the sediment microbial community through selection for different strains of microorganisms. The Pine River inoculum removed an average of 1.25 bromines per biphenyl molecule during a 32-week incubation period, resulting in a mixture potentially more accessible to aerobic degradation processes. No ortho bromine removal was observed. However, when Firemaster was incubated with Hudson River microorganisms that had been repeatedly transferred on a pyruvate medium amended with Aroclor 1242, 17% of the meta and para bromines were removed after 16 weeks of incubation and additional debromination products, including 2-bromobiphenyl and biphenyl, were detected. This suggests the possibility for ortho debromination, since all components of the Firemaster mixture have at least one ortho-substituted bromine.(ABSTRACT TRUNCATED AT 250 WORDS)     

The use of compound heterozygotes and Hprt selection to analyze X-linked mottled alleles associated with prenatal lethality

X-linked mutant alleles associated with prenatal male lethality are difficult to analyze because only heterozygous females are readily available for study. Genomic analysis of the mutant allele is facilitated by the construction of somatic cell hybrids because this enables the segregation of the X Chromosomes (Chrs) that carry the mutant and wild-type alleles. We describe here a method that ensures that the X Chr carrying the mutant allele is retained in somatic cell hybrids in an active selectable state. This is achieved by mating heterozygous females to males that carry a mutation at the hypoxanthine phosphoribosyl transferase (Hprt) locus. The resultant F₁ females are compound heterozygotes, and when cells from these females are fused to HPRT− Chinese hamster cells and subjected to selection in HAT medium, the only survivors are those hybrid cells that retain an active X Chr carrying the mutant allele together with the wild-type Hprt allele. We use hybrids constructed by this method to demonstrate that there are no gross deletions or genomic rearrangements present in three mottled alleles associated with prenatal male lethality. Received: 8 January 1996 / Accepted: 29 February 1996     

Diffusion Boundary Layers Ameliorate the Negative Effects of Ocean Acidification on the Temperate Coralline Macroalga Arthrocardia corymbosa

Anthropogenically-modulated reductions in pH, termed ocean acidification, could pose a major threat to the physiological performance, stocks, and biodiversity of calcifiers and may devalue their ecosystem services. Recent debate has focussed on the need to develop approaches to arrest the potential negative impacts of ocean acidification on ecosystems dominated by calcareous organisms. In this study, we demonstrate the role of a discrete (i.e. diffusion) boundary layer (DBL), formed at the surface of some calcifying species under slow flows, in buffering them from the corrosive effects of low pH seawater. The coralline macroalga Arthrocardia corymbosa was grown in a multifactorial experiment with two mean pH levels (8.05 ‘ambient’ and 7.65 a worst case ‘ocean acidification’ scenario projected for 2100), each with two levels of seawater flow (fast and slow, i.e. DBL thin or thick). Coralline algae grown under slow flows with thick DBLs (i.e., unstirred with regular replenishment of seawater to their surface) maintained net growth and calcification at pH 7.65 whereas those in higher flows with thin DBLs had net dissolution. Growth under ambient seawater pH (8.05) was not significantly different in thin and thick DBL treatments. No other measured diagnostic (recruit sizes and numbers, photosynthetic metrics, %C, %N, %MgCO₃) responded to the effects of reduced seawater pH. Thus, flow conditions that promote the formation of thick DBLs, may enhance the subsistence of calcifiers by creating localised hydrodynamic conditions where metabolic activity ameliorates the negative impacts of ocean acidification.     

Somatostatin inhibits insulin secretion by a G-protein-mediated decrease in Ca2+ entry through voltage-dependent Ca2+ channels in the beta cell.

We tested the hypothesis that somatostatin (SRIF) inhibits insulin secretion from an SV40 transformed hamster beta cell line (HIT cells) by an effect on the voltage-dependent Ca2+ channels and examined whether G-proteins were involved in the process. Ca2+ currents were recorded by the whole cell patch-clamp method, the free cytosolic calcium, [Ca2+]i, was monitored in HIT cells by fura-2, and cAMP and insulin secretion were measured by radioimmunoassay. SRIF decreased Ca2+ currents, [Ca2+]i, and basal insulin secretion in a dose-dependent manner over the range of 10(-12)-10(-7)M. The increase in [Ca2+]i and insulin secretion induced by either depolarization with K+ (15 mM) or by the Ca2+ channel agonist, Bay K 8644 (1 microM) was attenuated by SRIF in a dose-dependent manner over the same range of 10(-12)-10(-7) M. the half-maximal inhibitory concentrations (IC50) for SRIF inhibition of insulin secretion were 8.6 X 10(-12) M and 8.3 X 10(-11) M for K+ and Bay K 8644-stimulated secretion and 1 X 10(-10) M and 2.9 X 10(-10) M for the SRIF inhibition of the K+ and Bay K 8644-induced rise in [Ca2+]i, respectively. SRIF also attenuated the rise in [Ca2+]i induced by the cAMP-elevating agent, isobutylmethylxanthine (1 mM) in the presence of glucose. Bay K 8644, K+ and SRIF had no significant effects on cAMP levels and SRIF had no effects on adenylyl cyclase activity at concentrations lower than 1 microM. SRIF (100 nM) did not change K+ efflux (measured by 86Rb+) through ATP-sensitive K+ channels in HIT cells. SRIF (up to 1 microM) had no significant effect on membrane potential measured by bisoxonol fluorescence. Pretreatment of the HIT cells with pertussis toxin (0.1 microgram/ml) overnight abolished the effects of SRIF on Ca2+ currents, [Ca2+]i and insulin secretion implying a G-protein dependence in SRIF's actions. Thus, one mechanism by which SRIF decreases insulin secretion is by inhibiting Ca2+ influx through voltage-dependent Ca2+ channels, an action mediated through a pertussis toxin-sensitive G-protein.     

Bovine pituitary intraglandular colloid: a transport medium

Bovine pituitary intraglandular colloid is formed by the cyclic degeneration of marginal cells lining the intermediate lobe and is housed in the intraglandular lumen (residual lumen). The lumen communicates with the subarachnoid cerebrospinal fluid space by well defined channels. Electrophoresis in acrylamide gel shows bovine pituitary intraglandular colloid as having double protein bands identical to the protein in bovine and human cerebrospinal fluid. These studies demonstrate two distinct bands in the gamma region for colloid, not apparent in the normal bovine or human cerebrospinal fluid due to the low concentration of gamma globulins. We conclude that pituitary colloid, laden with immunoreactive fragments of various pituitary hormones, is discharged from the hypophyseal intraglandular space, directly into the subarachnoid cerebrospinal fluid space.