Yeast mitochondria (Saccharomyces cerevisiae) contain Ca(2+)-independent phospholipase A1 and A2 activities: effect of respiratory state

We demonstrate that both phospholipase A1 and phospholipase A2 are associated with isolated yeast mitochondria (Saccharomyces cerevisiae). Activity assays indicate that, unlike most other mitochondrial phospholipases A, the yeast enzymes are Ca(2+)-independent with acidic (pH 4-5) as well as alkaline (pH 8-9) pH optima. Data obtained with mitochondria isolated from either fermenting or respiring cells, and initial observations with a petite strain, strongly suggest that a phospholipase A2 with an acidic pH optimum functions in the in vivo adaptation and maintenance of mitochondrial membranes required for respiration.     

Prediction of Heterosis from DNA Fingerprints in Chickens

To assess the value of DNA fingerprints for the prediction of heterosis in chickens, retrospective analyses of data from three crossbreeding experiments and DNA fingerprints (DFP) of parental strains were conducted using two minisatellite and one middle-repetitive DNA probes. DFP bands were assessed on pooled DNA samples of 10-15 individuals per parental genetic group. The number of DFP bands evaluated in the experiments ranged from 81 to 139. The probes varied in their predictive value, but predictability of heterosis generally increased with multiple probes. Highly significant correlations (0.68-0.87) between band sharing ratios (SH) and heterosis were found in 25 crosses of White Leghorns in the first egg production cycle for age at sexual maturity, egg production, and mature body weight: traits with heterosis of 10% or more of the means. Regressions of SH explained 78.4% of the variation in heterosis in age at sexual maturity, 60.2% in egg production and 46.4% in mature body weight. For ``broiler' traits with heterosis of <1%, none of the correlations, based on 13 crosses, were significant. It was concluded that multilocus probe DFP of pooled DNA samples show promise as predictors of heterosis.     

A major difference between the divergence patterns within the lines-1 families in mice and voles

L1 retroposons are represented in mice by subfamilies of interspersed sequences of varied abundance. Previous analyses have indicated that subfamilies are generated by duplicative transposition of a small number of members of the L1 family, the progeny of which then become a major component of the murine L1 population, and are not due to any active processes generating homology within preexisting groups of elements in a particular species. In mice, more than a third of the L1 elements belong to a clade that became active approximately 5 Mya and whose elements are > or = 95% identical. We have collected sequence information from 13 L1 elements isolated from two species of voles (Rodentia: Microtinae: Microtus and Arvicola) and have found that divergence within the vole L1 population is quite different from that in mice, in that there is no abundant subfamily of homologous elements. Individual L1 elements from voles are very divergent from one another and belong to a clade that began a period of elevated duplicative transposition approximately 13 Mya. Sequence analyses of portions of these divergent L1 elements (approximately 250 bp each) gave no evidence for concerted evolution having acted on the vole L1 elements since the split of the two vole lineages approximately 3.5 Mya; that is, the observed interspecific divergence (6.7%-24.7%) is not larger than the intraspecific divergence (7.9%-27.2%), and phylogenetic analyses showed no clustering into Arvicola and Microtus clades.      

Molecular phylogeny and divergence times of drosophilid species

The phylogenetic relationships and divergence times of 39 drosophilid species were studied by using the coding region of the Adh gene. Four genera--Scaptodrosophila, Zaprionus, Drosophila, and Scaptomyza (from Hawaii)--and three Drosophila subgenera--Drosophila, Engiscaptomyza, and Sophophora--were included. After conducting statistical analyses of the nucleotide sequences of the Adh, Adhr (Adh-related gene), and nuclear rRNA genes and a 905-bp segment of mitochondrial DNA, we used Scaptodrosophila as the outgroup. The phylogenetic tree obtained showed that the first major division of drosophilid species occurs between subgenus Sophophora (genus Drosophila) and the group including subgenera Drosophila and Engiscaptomyza plus the genera Zaprionus and Scaptomyza. Subgenus Sophophora is then divided into D. willistoni and the clade of D. obscura and D. melanogaster species groups. In the other major drosophilid group, Zaprionus first separates from the other species, and then D. immigrans leaves the remaining group of species. This remaining group then splits into the D. repleta group and the Hawaiian drosophilid cluster (Hawaiian Drosophila, Engiscaptomyza, and Scaptomyza). Engiscaptomyza and Scaptomyza are tightly clustered. Each of the D. repleta, D. obscura, and D. melanogaster groups is monophyletic. The splitting of subgenera Drosophila and Sophophora apparently occurred about 40 Mya, whereas the D. repleta group and the Hawaiian drosophilid cluster separated about 32 Mya. By contrast, the splitting of Engiscaptomyza and Scaptomyza occurred only about 11 Mya, suggesting that Scaptomyza experienced a rapid morphological evolution. The D. obscura and D. melanogaster groups apparently diverged about 25 Mya. Many of the D. repleta group species studied here have two functional Adh genes (Adh-1 and Adh-2), and these duplicated genes can be explained by two duplication events.      

Activity of a “thermostable exotoxin” of Bacillus thuringiensis subsp. morrisoni in the Salmonella/microsomal assay for bacterial mutagenicity

The purpose of this study was to employ the Salmonella/microsomal assay (Ames test) to investigate the mutagenic potential of a thermostable exotoxin of Bacillus thuringiensis subsp. morrisoni. Bacteria are ideal for the detection of infrequently occurring point mutations because the large number of organisms (200 to 400 million bacteria per plate) exposed to the mutagen at any one time increases the possibility of observing a random mutational event. The exotoxin used in this study was produced using the shaker flask fermentation procedure with mineral casein broth. A Petri dish method of bioassay using fresh bovine feces was used to determine the efficacy of the exotoxin against horn flies. The LD₅₀ was found to be 5.35 μl/g of feces. Five bacterial tester strains were identified and characterized for the genetic markers described by Ames et al. (B. N. Ames et al., 1975, Mutat. Res., 31, 347–364). Appropriate doses of the B. thuringiensis supernatant, solvent or positive control were added to agar plates. The supernatant was tested at five dose levels against all five strains of bacteria. Controls of bacteria only were included for spontaneous reversions. All treatments were performed in triplicate. The numbers of revertant colonies from each set of triplicate plates were averaged and the standard deviation calculated and compared to that found with the solvent control. The negative controls, positive controls, and sterility controls all fulfilled requirements for determination of a valid test. No detectable mutagenic activity was found for the thermostable exotoxin of B. thuringiensis morrisoni.     

Sulfate reduction and S-oxidation in a moorland pool sediment

In an oligotrophic moorland pool in The Netherlands, S cycling near the sediment/water boundary was investigated by measuring (1) SO₄ ^2– reduction rates in the sediment, (2) depletion of SO₄ ^2– in the overlying water column and (3) release of³⁵S from the sediment into the water column. Two locations differing in sediment type (highly organic and sandy) were compared, with respect to reduction rates and depletion of SO₄ ^2– in the overlying water.Sulfate reduction rates in sediments of an oligotrophic moorland pool were estimated by diagenetic modelling and whole core³⁵SO₄ ^2– injection. Rates of SO₄ ^2– consumption in the overlying water were estimated by changes in SO₄ ^2– concentration over time in in situ enclosures. Reduction rates ranged from 0.27–11.2 mmol m^–2 d^–1. Rates of SO₄ ^2– uptake from the enclosed water column varied from –0.5, –0.3 mmol m^–2 d^–1 (November) to 0.43–1.81 mmol m^–2 d^–1 (July, August and April). Maximum rates of oxidation to SO₄ ^2– in July 1990 estimated by combination of SO₄ ^2– reduction rates and rates of in situ SO₄ ^2– uptake in the enclosed water column were 10.3 and 10.5 mmol m^–2 d^–1 at an organic rich and at a sandy site respectively.Experiments with³⁵S^2– and³⁵SO₄ ^2– tracer suggested (1) a rapid formation of organically bound S from dissimilatory reduced SO₄ ^2– and (2) the presence of mainly non SO₄ ^2–-S derived from reduced S transported from the sediment into the overlying water. A³⁵S^2– tracer experiment showed that about 7% of³⁵S^2– injected at 1 cm depth in a sediment core was recovered in the overlying water column.Sulfate reduction rates in sediments with higher volumetric mass fraction of organic matter did not significantly differ from those in sediments with a lower mass fraction of organic matter.Corresponding author     

A hybrid Azotobacter vinelandii-Clostridium pasteurianum nitrogenase iron protein that has in vivo and in vitro catalytic activity

Site-directed mutagenesis and gene replacement procedures were used to construct a mutant strain of Azotobacter vinelandii which expresses a hybrid nitrogenase Fe protein. This hybrid Fe protein has its carboxyl-terminal 18 residues replaced with the 5 analogous residues from the Clostridium pasteurianum Fe protein sequence. The hybrid Fe protein is 13 amino acids smaller than the wild-type A. vinelandii Fe protein and has a net loss of 4 negatively charged residues, resulting in a change in size and charge. The strain which produces the hybrid Fe protein remained capable of diazotrophic growth, albeit at a reduced rate. Also, the purified hybrid Fe protein exhibited a maximum activity about one-half that of native Fe protein. These results demonstrate that the tight, inactive complex which is formed when A. vinelandii MoFe protein and C. pasteurianum Fe protein are mixed in heterologous reconstitution experiments cannot be accounted for only by differences in the A. vinelandii and C. pasteurianum Fe protein primary sequences located at their respective carboxyl termini.