The genetic basis of alcoholism: multiple phenotypes,many genes,complex networks

Alcoholism is a significant public health problem. A picture of the genetic architecture underlying alcohol-related phenotypes is emerging from genome-wide association studies and work on genetically tractable model organisms.     

Modulation of Escherichia coli DNA methyltransferase activity by biologically derived GATC-flanking sequences

Escherichia coli DNA adenine methyltransferase (EcoDam) methylates the N-6 position of the adenine in the sequence 5'-GATC-3' and plays vital roles in gene regulation, mismatch repair, and DNA replication. It remains unclear how the small number of critical GATC sites involved in the regulation of replication and gene expression are differentially methylated, whereas the approximately 20,000 GATCs important for mismatch repair and dispersed throughout the genome are extensively methylated. Our prior work, limited to the pap regulon, showed that methylation efficiency is controlled by sequences immediately flanking the GATC sites. We extend these studies to include GATC sites involved in diverse gene regulatory and DNA replication pathways as well as sites previously shown to undergo differential in vivo methylation but whose function remains to be assigned. EcoDam shows no change in affinity with variations in flanking sequences derived from these sources, but methylation kinetics varied 12-fold. A-tracts immediately adjacent to the GATC site contribute significantly to these differences in methylation kinetics. Interestingly, only when the poly(A) is located 5' of the GATC are the changes in methylation kinetics revealed. Preferential methylation is obscured when two GATC sites are positioned on the same DNA molecule, unless both sites are surrounded by large amounts of nonspecific DNA. Thus, facilitated diffusion and sequences immediately flanking target sites contribute to higher order specificity for EcoDam; we suggest that the diverse biological roles of the enzyme are in part regulated by these two factors, which may be important for other enzymes that sequence-specifically modify DNA.     

Geomicrobial characterization of gas hydrate-bearing sediments along the mid-Chilean margin

Bacterial diversity in eight sediment cores from the mid-Chilean margin was studied using length heterogeneity (LH)-PCR, and described in relation to in situ geochemical conditions. DNA from the sulfate-methane transition (SMT) of three cores [one containing methane gas; two proximal to a gas hydrate mound (GHM)] was cloned and sequenced. Clones related to uncultured relatives of Desulfosarcina variabilis were found in all clone libraries and dominated one. Desulfosarcina variabilis related clones were similar to phylotypes observed at the SMT in association with anaerobic methane oxidation in the Eel River basin, Cascadia margin and the Gulf of Mexico. The LH-PCR amplicon associated with D. variabilis clones matched the amplicon that dominated most SMT samples, indicating environmental selection for D. variabilis relatives. Clones related to the Verrucomicrobia dominated the library for the methane gas-containing core. Uncultured Treponema relatives dominated the library for the core obtained on the edge of a GHM. Statistical analysis using geochemical data to describe variance in LH-PCR data revealed that stable carbon isotope ratios of dissolved inorganic carbon are the principal structuring factor on SMT communities. These data suggest that D. variabilis relatives are involved in anaerobic oxidation of methane at the SMT in Chilean margin sediments.     

Oncogenicity of DNA in vivo: Tumor induction with expression plasmids for activated H-ras and c-myc

All vaccines and other biological products contain contaminating residual DNA derived from the production cell substrate. Whether this residual cell-substrate DNA can induce tumors in vaccine recipients and thus represent a risk factor has been debated for over 50 years without resolution. As a first step in resolving this issue, we have generated expression plasmids for the activated human H-ras oncogene and for the murine c-myc proto-oncogene. Their oncogenic activity was confirmed in vitro using the focus-formation transformation assay. Two strains of adult and newborn immune-competent mice were inoculated with different amounts of either plasmid alone or with a combination of the H-ras and c-myc plasmids. Tumors developed only in mice inoculated with both plasmids and only at the highest amount of DNA (12.5 microg of each plasmid). The NIH Swiss mouse was more sensitive than the C57BL/6 mouse, and newborn animals were more sensitive than adults. Cell lines were established from the tumors. PCR and Southern hybridization analyses demonstrated that both inoculated oncogenes were present in all of the tumor-derived cell lines and that the cells in the tumors were clonal. Western analysis demonstrated that both oncoproteins were expressed in these cell lines. These results demonstrate that cellular oncogenes can induce tumors following subcutaneous inoculation. Such information provides a possible way of evaluating and estimating the theoretical oncogenic risk posed by residual cell-substrate DNA in vaccines.     

Host-retrovirus arms race: trimming the budget

In this issue of Cell Host & Microbe, OhAinle et al., 2008 report that APOBEC3H, a potent innate retroviral restriction factor in primates, lost its function twice independently during recent evolution in humans, stressing an ever present trade-off between benefit and cost of protection against pathogens.     

Stable carbon isotope analysis of nucleic acids to trace sources of dissolved substrates used by estuarine bacteria.

The natural abundance of stable carbon isotopes measured in bacterial nucleic acids extracted from estuarine bacterial concentrates was used to trace sources of organic matter for bacteria in aquatic environments. The stable carbon isotope ratios of Pseudomonas aeruginosa and nucleic acids extracted from cultures resembled those of the carbon source on which bacteria were grown. The carbon isotope discrimination between the substrate and total cell carbon from bacterial cultures averaged 2.3% +/- 0.6% (n = 13). Furthermore, the isotope discrimination between the substrate and nucleic acids extracted from bacterial cultures was 2.4% +/- 0.4% (n = 10), not significantly different from the discrimination between bacteria and the substrate. Estuarine water samples were prefiltered through 1-micron-pore-size cartridge filters. Bacterium-sized particles in the filtrates were concentrated with tangential-flow filtration and centrifugation, and nucleic acids were then extracted from these concentrates. Hybridization with 16S rRNA probes showed that approximately 90% of the nucleic acids extracted on two sample dates were of eubacterial origin. Bacteria and nucleic acids from incubation experiments using estuarine water samples enriched with dissolved organic matter from Spartina alterniflora and Cyclotella caspia had stable carbon isotope values similar to those of the substrate sources. In a survey that compared diverse estuarine environments, stable carbon isotopes of bacteria grown in incubation experiments ranged from -31.9 to -20.5%. The range in isotope values of nucleic acids extracted from indigenous bacteria from the same waters was similar, -27.9 to -20.2%. Generally, the lack of isotope discrimination between bacteria and nucleic acids that was noted in the laboratory was observed in the field.(ABSTRACT TRUNCATED AT 250 WORDS)     

Ecological responses of dominant grasses along two climatic gradients in the Great Plains of the United States

Abstract. Few empirical data exist to examine the influence of regional scale environmental gradients on productivity patterns of plant species. In this paper we analyzed the productivity of several dominant grass species along two climatic gradients, mean annual precipitation (MAP) and mean annual temperature (MAT), in the Great Plains of the United States. We used climatic data from 296 weather stations, species production data from Natural Resource Conservation Service rangeland surveys and a geographic information system to spatially integrate the data. Both MAP and MAT were significantly related to annual above-ground net primary production (ANPP). MAP explained 54% to 89% of the variation in ANPP of two C₄ short-grasses, Bouteloua gracilis and Buchloë dactyloides, and two C₄ tall-grasses, Andropogon gerardii and Schizachyrium scoparium (= Andropogon scoparius). MAT explained 19% to 41% of the variation in ANPP of two C₄ grasses, B. gracilis and B. dactyloides, and 41% to 66% of the variation in ANPP of two C₃ grasses, Agropyron smithii and Stipa comata. ANPP patterns for species along both gradients were described by either linear, negative exponential, logistic, normal or skewed curves. Patterns of absolute ANPP (g/m²) for species differed from those of relative ANPP (%) along the MAP gradient. Responses were similar for species with common functional characteristics (e.g. short-grasses, tall-grasses, C₃, C₄). Our empirical results support asymmetric responses of species to environmental gradients. Results demonstrate the importance of species attributes, type of environmental gradient and measure of species importance (relative or absolute productivity) in evaluating ecological response patterns.