A Comprehensive Study of Genic Variation in Natural Populations of Drosophila melanogaster. III. Variations in Genetic Structure and Their Causes between Drosophila melanogaster and Its Sibling Species Drosophila simulans

The natural populations of Drosophila melanogaster and Drosophila simulans were compared for their genetic structure. A total of 114 gene-protein loci were studied in four mainland (from Europe and Africa) and an island (Seychelle) populations of D. simulans and the results were compared with those obtained on the same set of homologous loci in fifteen worldwide populations of D. melanogaster. The main results are as follows: (1) D. melanogaster shows a significantly higher proportion of loci polymorphic than D. simulans (52% vs. 39%, P<0.05), (2) both species have similar mean heterozygosity and mean number of alleles per locus, (3) the two species share some highly polymorphic loci but they do not share loci that show high geographic differentiation, and (4) D. simulans shows significantly less geographic differentiation than D. melanogaster. The differences in genetic differentiation between the two species are limited to loci located on the X and second chromosomes only; loci on the third chromosome show similar level of geographic differentiation in both species. These two species have previously been shown to differ in their pattern of variation for chromosomal polymorphisms, quantitative and physiological characters, two-dimensional electrophoretic (2DE) proteins, middle repetitive DNA and mitochondrial DNA. Variation in niche-widths and/or genetic "strategies" of adaptation appear to be the main causes of differences in the genetic structure of these two species.     

The consequences of fluctuating selection for isozyme polymorphisms in Daphnia

Temporal sequences of allele frequencies in natural populations of Daphnia are analyzed to obtain the mean and variance of the selection coefficient for both asexual and sexual phases. In general, the alleles at enzyme loci appear to be quasi-neutral. Although significant variation exists for the estimated selection coefficients, the means are in all cases close to zero. Estimates of the variance of selection intensity are applied to existing models to demonstrate the implications of fluctuating selection for the spatial and temporal distribution of gene frequencies in Daphnia. The empirical and analytical results are shown to provide a possible solution to some previously puzzling aspects of Daphnia population genetic surveys. Neither genetic drift nor diversifying selection are necessary conditions for the local diversification of gene frequencies.     

Three-dimensional representation of chemical gradients

Perspective display techniques are applied to chemical and biochemical data sets. These represent spatially distributed gradients of reactive compounds that participate in pattern-formation processes due to reaction-diffusion or reaction-convection coupling. The patterns form in thin solution layers and are observed as chemical waves in the Belousov-Zhabotinskii reaction, as convection-induced stationary structures during oscillating glycolysis in yeast cytoplasm, and as the diffusive spreading of enzyme-catalyzed metabolic turnover in a substrate layer. The digital data are measured with a two-dimensional spectrophotometer based on a computerized video equipment with high spatial, temporal and intensity resolution. By application of three-dimensional procedures detailed structural properties of chemical and biochemical model systems will be presented yielding localization of reaction and transport events.     

Low Temperature-Induced Decrease in trans-Delta-Hexadecenoic Acid Content Is Correlated with Freezing Tolerance in Cereals

The effect of growth at 5°C on the trans-Δ³-hexadecenoic acid content of phosphatidyl(d)glycerol was examined in a total of eight cultivars of rye (Secale cereale L.) and what (Triticum aestivum L.) of varying freezing tolerance. In these monocots, low temperature growth caused decreases in the trans-Δ³-hexadecenoic acid content of between 0 and 74% with concomitant increases in the palmitic acid content of phosphatidyl(d)glycerol. These trends were observed for whole leaf extracts as well as isolated thylakoids. The low growth temperature-induced decrease in the trans-Δ³-hexadecenoic acid content was shown to be a linear function (r² = 0.954) of freezing tolerance in these cultivars. Of the six cold tolerant dicotyledonous species examined, only Brassica and Arabidopsis thaliana L. cv Columbia exhibited a 42% and 65% decrease, respectively, in trans-Δ³-hexadecenoic acid content. Thus, the relationship between the change in trans-Δ³-hexadecenoic acid content of phosphatidyl(d)glycerol and freezing tolerance cannot be considered a general one for all cold tolerant plant species. However, species which exhibited a low growth temperature-induced decrease in trans-Δ³-hexadecenoic acid also exhibited a concomitant shift in the in vitro organization of the light harvesting complex II from a predominantly oligomeric form to the monomeric form. We conclude that the proposed role of phosphatidyl(d)glycerol in modulating the organization of light harvesting complex II as a function of growth temperature manifests itself to varying degrees in different plant species. A possible physiological role for this phenomenon with respect to low temperature acclimation and freezing tolerance in cereals is discussed.     

Thermotolerance of isolated mitochondria associated with heat shock proteins

Mitochondria isolated from 2-day-old etiolated soybean (Glycine max) seedlings which had been subjected to various heat shock treatments, i.e. (A) 28°C (2 h), (B) 38°C (2 h), (C) 38°C (2 h)-42.5°C (0.5 h), and (D) 38°C (2 h)-42.5°C (0.5 h)-28°C (4 h), were monitored for O₂ uptake using an oxygen electrode. Mitochondria isolated after all four heat shock treatments were active in O₂ consumption at 28°C in response to succinate and ADP (derived P/O ratios were 1.6, 1.7, 1.3, and 1.3, respectively.) The mitochondria from all four treatments were also active in O₂ uptake at 42.5°C. However, only mitochondria isolated after treatment (C) were tightly coupling at 42.5°C (derived ADP/O ratio was about 1.4). Combined with our earlier findings on the subcellular localization of heat shock proteins, our present data demonstrate that association of heat shock proteins with mitochondria by treatment (C) enables them to phosphorylate at 42.5°C (i.e. they become thermotolerant). Isolated mitochondria from treatment (C) and treatment (A) were compared by electron microscopy. They appeared to be very similar and no significant ultrastructural differences were noted.     

Evidence for Chloroplastic Succinate Dehydrogenase Participating in the Chloroplastic Respiratory and Photosynthetic Electron Transport Chains of Chlamydomonas reinhardtii

A method for isolating intact chloroplasts from Chlamydomonas reinhardtii F-60 was developed from the Klein, Chen, Gibbs, Platt-Aloia procedure ([1983] Plant Physiol 72: 481-487). Protoplasts, generated by treatment with autolysine, were lysed with a solution of digitonin and fractionated on Percoll step gradients. The chloroplasts were assessed to be 90% intact (ferricyanide assay) and free from cytoplasmic contamination (NADP isocitrate dehydrogenase activity) and to range from 2 to 5% in mitochondrial contamination (cytochrome c oxidase activity). About 25% of the cellular succinate dehydrogenase activity (21.6 micromoles per milligram chlorophyll per hour, as determined enzymically) was placed within the chloroplast. Chloroplastic succinate dehydrogenase had a K_m for succinate of 0.55 millimolar and was associated with the thylakoidal material derived from the intact chloroplasts. This same thylakoidal material, with an enzymic assay of 21.6 micromoles per milligram chlorophyll per hour was able to initiate a light-dependent uptake of oxygen at a rate of 16.4 micromoles per milligram chlorophyll per hour when supplied with succinate and methyl viologen. Malonate was an apparent competitive inhibitor of this reaction. The succinate dehydrogenase activity present in the chloroplast was sufficient to account for the photoanaerobic rate of acetate dissimilation in H₂ adapted Chlamydomonas (M Gibbs, RP Gfeller, C Chen [1986] Plant Physiol 82: 160-166).     

Properties of a Lipoxygenase in Green Algae (Oscillatoria sp.)

A lipoxygenase preparation was obtained from green algae Oscillatoria sp. and was shown to differ from previous described lipoxygenases in the positional specificity and pH characteristics of the dioxygenation reaction. The enzyme had a pH optimum at 8.8 and was inactive at pH 6. Oscillatoria lipoxygenase converted linoleic acid into two products: 13-hydroperoxylinoleic acid (52%) and 9-hydroperoxylinoleic acid (48%). The molecular weight of the enzyme was estimated at 124,000. Esculetin was found to be the best inhibitor of the enzyme activity.     

Protoplast volume:water potential relationship and bound water fraction in spinach leaves

Methods used to estimate the (nonosmotic) bound water fraction (BWF) (i.e. apoplast water) of spinach (Spinacia oleracea L.) leaves were evaluated. Studies using three different methods of pressure/volume (P/V) curve construction all resulted in a similar calculation of BWF; approximately 40%. The theoretically derived BWF, and the water potential (Ψ_w)/relative water content relationship established from P/V curves were used to establish the relationship between protoplast (i.e. symplast) volume and Ψ_w. Another method of establishing the protoplast volume/Ψ_w relationship in spinach leaves was compared with the results from P/V curve experiments. This second technique involved the vacuum infiltration of solutions at a range of osmotic potentials into discs cut from spinach leaves. These solutions contained radioactively labeled H₂O and sorbitol. This dual label infiltration technique allowed for simultaneous measurement of the total and apoplast volumes in leaf tissue; the difference yielded the protoplast volume. The dual label infiltration experiments and the P/V curve constructions both showed that below −1 megapascals, protoplast volume decreases sharply with decreasing water potential; with 50% reduction in protoplast volume occurring at −1.8 megapascals leaf water potential.     

Tungstate, a molybdate analog inactivating nitrate reductase, deregulates the expression of the nitrate reductase structural gene

Nitrate reductase (NR, EC 1.6.6.1) from higher plants is a homodimeric enzyme carrying a molybdenum cofactor at the catalytic site. Tungsten can be substituted for molybdenum in the cofactor structure, resulting in an inactive enzyme. When nitratefed Nicotiana tabacum plants were grown on a nutrient solution in which tungstate was substituted for molybdate, NR activity in the leaves decreased to a very low level within 24 hours while NR protein accumulated progressively to a level severalfold higher than the control after 6 days. NR mRNA level in molybdate-grown plants exhibited a considerable day-night fluctuation. However, when plants were treated with tungstate, NR mRNA level remained very high. NR activity and protein increased over a 24-hour period when nitrate was added back to N-starved molybdate-grown plants. NR mRNA level increased markedly during the first 2 hours and then decreased. In the presence of tungstate, however, the induction of NR activity by nitrate was totally abolished while high levels of NR protein and mRNA were both induced, and the high level of NR mRNA was maintained over a 10-hour period. These results suggest that the substitution of tungsten for molybdenum in NR complex leads to an overexpression of the NR structural gene. Possible mechanisms involved in this deregulation are discussed.