A single nucleotide polymorphism in the<Emphasis Type="Italic"> Emp3</Emphasis> gene defines the H4 minor histocompatibility antigen

Minor histocompatibility antigens (minor H antigen) elicit strong T-cell-mediated responses during both graft rejection and graft versus leukemia (GvL) among MHC-matched individuals (where MHC is major histocompatibility complex). Employing expression-cloning methodology, we have identified a cDNA clone, MI-35, encoding the immunodominant H4^b minor H antigen within the classical mouse H4 complex. The minimal antigenic epitope derived from H4^b presented on K^b class I MHC is SGIVYIHL (SYL8) and the polymorphism is due to CT nucleotide modification in p3 resulting in the change of threonine (ACT) to isoleucine (ATT). The results presented here demonstrate that amino acid variation in the allelic epitopes results in the low abundance of H4^a peptide. The differential peptide copy number resulted in an immunodominant cytotoxic T cells (CTL) response directed against H4^b while the anti-B6 response directed against H4^a was easily dominated. These results provide a molecular mechanism for the H4 minor H antigen and suggest a novel mechanism by which alloantigenic disparity caused by conservative amino acid changes can be augmented by posttranslational antigen processing events.     

Disinfection of bacterial biofilms in pilot-scale cooling tower systems

The impact of continuous chlorination and periodic glutaraldehyde treatment on planktonic and biofilm microbial communities was evaluated in pilot-scale cooling towers operated continuously for 3 months. The system was operated at a flow rate of 10,080 l day^?1. Experiments were performed with a well-defined microbial consortium containing three heterotrophic bacteria: Pseudomonas aeruginosa, Klebsiella pneumoniae and Flavobacterium sp. The persistence of each species was monitored in the recirculating cooling water loop and in biofilms on steel and PVC coupons in the cooling tower basin. The observed bacterial colonization in cooling towers did not follow trends in growth rates observed under batch conditions and, instead, reflected differences in the ability of each organism to remain attached and form biofilms under the high-through flow conditions in cooling towers. Flavobacterium was the dominant organism in the community, while P. aeruginosa and K. pneumoniae did not attach well to either PVC or steel coupons in cooling towers and were not able to persist in biofilms. As a result, the much greater ability of Flavobacterium to adhere to surfaces protected it from disinfection, whereas P. aeruginosa and K. pneumoniae were subject to rapid disinfection in the planktonic state.     

CYP3A4-V and prostate cancer in African Americans: causal or confounding association because of population stratification?

CYP3A4-V, an A to G promoter variant associated with prostate cancer in African Americans, exhibits large differences in allele frequency between populations. Given that the African American population is genetically heterogeneous because of its African ancestry and subsequent admixture with European Americans, case-control studies with African Americans are highly susceptible to spurious associations. To test for association with prostate cancer, we genotyped CYP3A4-V in 1376 (2 N) chromosomes from prostate cancer patients and age- and ethnicity-matched controls representing African Americans, Nigerians, and European Americans. To detect population stratification among the African American samples, 10 unlinked genetic markers were genotyped. To correct for the stratification, the uncorrected association statistic was divided by the average of association statistics across the 10 unlinked markers. Sharp differences in CYP3A4-V frequencies were observed between Nigerian and European American controls (0.87 and 0.10, respectively; P<0.0001). African Americans were intermediate (0.66). An association uncorrected for stratification was observed between CYP3A4-V and prostate cancer in African Americans (P=0.007). A nominal association was also observed among European Americans (P=0.02) but not Nigerians. In addition, the unlinked genetic marker test provided strong evidence of population stratification among African Americans. Because of the high level of stratification, the corrected P-value was not significant (P=0.25). Follow-up studies on a larger dataset will be needed to confirm whether the association is indeed spurious; however, these results reveal the potential for confounding of association studies by using African Americans and the need for study designs that take into account substructure caused by differences in ancestral proportions between cases and controls.     

Antioxidant enzymes, free-radical damage, and response to paraquat in liver and kidney of long-living growth hormone receptor/binding protein gene-disrupted mice.

Numerous studies have shown that the lifespan can be extended by caloric restriction or by altering the growth hormone (GH)-insulin-like growth factor 1 signaling pathway. Both of these manipulations produce physiological alterations, such as increased insulin sensitivity, and reduced glucose levels and body size. However, it is difficult to evaluate whether these are merely correlates of delayed aging or whether they have a direct causal effect on lifespan. One parameter that has been demonstrated to have causal, positive effects on longevity in invertebrates is improved antioxidant defenses. We measured activities of antioxidant enzymes Cu/Zn superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx) and quantified free-radical damage by lipid peroxidation (LP) and protein oxidation (PO) measurements in liver and kidney tissues, and evaluated the response to paraquat-induced oxygen toxicity in the long-living GH receptor/binding protein gene knockout (GHR-KO) mouse. We found that in the kidney, SOD was lower and GPx was higher in GHR-KO mice, and LP was higher in female GHR-KO mice only. In the liver, female GHR-KO mice had lower GPx, while male GHR-KO mice had lower CAT and higher LP. GHR-KO males were also more susceptible to paraquat toxicity compared to females or normal males. We conclude that in long-living GHR-KO mice, GH-resistance does not confer longevity by improved free-radical scavenging in the liver and kidney, suggesting that greater free-radical defenses in other tissues, or altered glucose metabolism may have a more central role in extending the lifespan of these animals.     

ISOLATION OF ALGAL GENES BY FUNCTIONAL COMPLEMENTATION OF YEAST1

Algal cDNAs were isolated and characterized by functional complementation of yeast auxotrophs. Two cDNA libraries, one derived from the diatom Phaeodactylum tricornutum Bohlin and the other from the dinoflagellate Crypthecodinium cohnii Biecheler, were constructed using the Saccharomyces cerevisiae expression vector pFL‐61. These libraries were used for functional complementation of auxotrophic markers in two yeast strains. Yeast tryptophan auxotrophs, complemented by the P. tricornutum library, contained a plasmid that encoded a two‐domain protein associated with tryptophan synthesis, indole‐3‐glycerol phosphate synthase‐N‐(5′‐phosphoribosyl) anthranilate isomerase. Another cDNA originating from the C. cohnii library rescued S. cervisiae from a defect in adenine biosynthesis. This cDNA encoded a fusion of phosphoribosylamidoimidazole‐succinocarboxamide synthetase and phosphoribosylaminoimidazole carboxylase, which correspond to the yeast ade1 and ade2 genes, respectively. These results demonstrate that heterologous functional complementation can be used to identify algal genes and may provide advantages over other gene discovery methods.     

Magnesium isotope fractionation during microbially enhanced forsterite dissolution

Bacillus subtilis endospore‐mediated forsterite dissolution experiments were performed to assess the effects of cell surface reactivity on Mg isotope fractionation during chemical weathering. Endospores present a unique opportunity to study the isolated impact of cell surface reactivity because they exhibit extremely low metabolic activity. In abiotic control assays, ²⁴Mg was preferentially released into solution during forsterite dissolution, producing an isotopically light liquid phase (δ²⁶Mg = ?0.39 ± 0.06 to ?0.26 ± 0.09‰) relative to the initial mineral composition (δ²⁶Mg = ?0.24 ± 0.03‰). The presence of endospores did not have an apparent effect on Mg isotope fractionation associated with the release of Mg from the solid into the aqueous phase. However, the endospore surfaces preferentially adsorbed ²⁴Mg from the dissolution products, which resulted in relatively heavy aqueous Mg isotope compositions. These aqueous Mg isotope compositions increased proportional to the fraction of dissolved Mg that was adsorbed, with the highest measured δ²⁶Mg (?0.08 ± 0.07‰) corresponding to the highest degree of adsorption (~76%). The Mg isotope composition of the adsorbed fraction was correspondingly light, at an average δ²⁶Mg of ?0.49‰. Secondary mineral precipitation and Mg adsorption onto secondary minerals had a minimal effect on Mg isotopes at these experimental conditions. Results demonstrate the isolated effects of cell surface reactivity on Mg isotope fractionation separate from other common biological processes, such as metabolism and organic acid production. With further study, Mg isotopes could be used to elucidate the role of the biosphere on Mg cycling in the environment.     

Comparing temporal patterns in body condition of ringed seals living within their core geographic range with those living at the edge

Ecological theory suggests that demographic responses by populations to environmental change vary depending on whether individuals inhabit central or peripheral regions within the species’ geographic range. Here, we tested this prediction by comparing a population of ringed seals Pusa hispida located at high latitudes as part of their core range (core) with a population located at the southern extremity of their range (peripheral). First, we compared the two regions’ environmental trends in timing of sea-ice breakup and freeze-up, open-water duration and the North Atlantic Oscillation (NAO). We found that the core region shifted to progressively warmer conditions in the early 1990s; whereas, in the peripheral region, the warming trend shifted in 1999 to one with no warming trend but high inter-annual variability. Next, we examined how body condition, inferred from blubber depth, responded to temporal changes in sea-ice and climatic variables – variables that have been shown to influence ringed seal demography. Core seals displayed minimal seasonal changes in body condition; whereas peripheral seals displayed a 20–60% amplitude seasonal change in body condition with a phase shift to earlier initiation of fat accumulation and loss. Finally, we tested for interannual differences and found that both core and peripheral seals responded similarly with decreased body condition following more positive NAO. Environmental variables influenced body condition in opposite directions between the two regions with core seals declining in body condition with later spring breakup and shorter open-water duration, whereas peripheral seals showed opposite relationships. Seals living at the core likely benefit from an evolved match between adaptation and environmental variation resulting in dampened seasonal and interannual fluctuations in body condition. Knowledge of how different populations respond to environmental change depending on geographic location within a species range can assist in anticipating population specific responses to climate warming.     

Mps1 promotes poleward chromosome movements in meiotic prometaphase

In prophase of meiosis I, homologous chromosomes pair and become connected by cross-overs. Chiasmata, the connections formed by cross-overs, enable the chromosome pair, called a bivalent, to attach as a single unit to the spindle. When the meiotic spindle forms in prometaphase, most bivalents are associated with one spindle pole and then go through a series of oscillations on the spindle, attaching to and detaching from microtubules until the partners of the bivalent become bioriented—attached to microtubules from opposite sides of the spindle. The conserved kinase, Mps1, is essential for the bivalents to be pulled by microtubules across the spindle in prometaphase. Here we show that MPS1 is needed for efficient triggering of the migration of microtubule-attached kinetochores toward the poles and promotes microtubule depolymerization. Our data support the model Mps1 acts at the kinetochore to coordinate the successful attachment of a microtubule and the triggering of microtubule depolymerization to then move the chromosome.