Darrell G. Yardley

Frequency-dependent viabilities of Drosophila pseudoobscura karyotypes

The viabilities between egg and adult life stages of Drosophila pseudoobscura karyotypes were studied at low, intermediate, and high frequencies. The viabilities of pairs of karyotypes were compared at each frequency and the viabilities of the three karyotypes, at one combination of frequencies. Eggs were counted into vials and samples taken of the adults emerging after viability selection. ST and CH gene arrangements of the third chromosome carrying different amylase alleles were used, and the karyotypes of adult flies were scored by gel electrophoresis. A statistical method related to the loglinear model was developed for estimating viabilities. This method takes account of the additional variability between replicates common in experiments of this kind and allows testing of nested hypotheses about the mechanism of selection. The viabilities of the homokaryotypes relative to the heterokaryotype were significantly higher at the low homokaryotypic frequency than at the higher ones. These viabilities do not show a consistent heterozygote advantage. This pattern of frequency-dependent viabilities will lead to a protected polymorphism for the gene arrangements, even in the absence of heterozygote advantage.

An amylase gene from Drosophila pseudoobscura is expressed in Escherichia coli: Functional selection and biochemical comparisons of the fly- and clone-produced amylases

An amylase gene from Drosophila pseudoobscura was isolated from a genomic library constructed in pBR322 and cloned in Escherichia coli by selecting for the ability of its product to hydrolyze starch, a carbon source not normally utilized by E. coli. Hybridization of pAMY17F to D. pseudoobscura polytene chromosomes shows a positive signal at the amylase pseudogene locus (bank 78, chromosome 3). The chimeric plasmid pAMY17F, has been altered in such a way as to increase amylase expression. Southern and Northern hybridizations to the cloned amylase DNA indicate that the source of the gene is from D. pseudoobscura. Biochemical properties such as pH optima, substrate specificities, electrophoretic analyses, inhibitor sensitivities, heat stabilities, temperature responsiveness and molecular weights indicate that the amylases produced by the fly and bacterial clone are similar and have similar properties. It appears that E. coli/pAMY17F is producing an amylase like that found in D. pseudoobscura.

The amylase system of Drosophila—I. Comparison of the amylases of D. pseudoobscura

Abstract 1. 1. Two amylases encoded by the Amy1.0 and Amy0.84 alleles of Drosophila pseudoobscura were compared in terms of pH optima, substrate specificities, inhibitor and activator sensitivities, and responses to temperature. 2. 2. Both crude extracts and partially purified preparations were studied. 3. 3. The amylases appeared quite similar to other α-amylases of Drosophila and possessed general propertes of α-amylases. 4. 4. A major difference between the two amylases was observed in terms of response to temperature: the Amy0.84 amylase appears to have a temperature optimum at 20°C whereas the Amy1.0 amylase shows increasing activity with increasing temperature over the 10–25°C range tested.

Effects of diet on amylase expression in the mosquitofish.

Diets high in various carbohydrates were fed to mosquitofish,Gambusia affinis holbrooki, to determine the effects on amylase expression. Both amylase activity and amount of amylase protein were used as measures of amylase expression. Fish were fed for 21 days in one experiment, seven days in a second experiment and 24 h in a third. The first experiment compared responses of fish fed on a high-starch diet relative to a control diet. The second and third experiments compared responses on four diets relative to the control diet: maltose, starch, glucose, and glucose + cyclic adenosine monophosphate (cAMP). In the first two experiments whole visceral extracts were used. In the third experiment, gut and hepatopancreatic extracts were examined separately. Diet had a significant affect on the amount of amylase in all three experiments but affected amylase activity only in the 24 h experiment. Generally, glucose decreased amylase expression while maltose or cAMP + glucose increased it. Length of feeding period and tissue type also had significant effects on amylase expression.

The amylase gene-enzyme system of chickens. II. Biochemical characterization of allozymes

The chicken amylase allozymes, AmyF and AmyS, were extracted from pancreatic tissues ofAmyF/FandAmyS/Sindividuals and purified. Activities were measured under various reaction conditions (=treatments) to assess whether the allozymes were functionally different. The amylases had properties typical of α-amylases, i.e., both were inhibited by ethylenediaminetetraacetate and α-amylase inhibitor from wheat, hadpH optima between 7.0 and 8.0, and could utilize a variety of substrates containing α 1,4 linkages. The amylases were also found to be inhibited by potassium phosphate buffer andp-chloromercuribenzoate. In terms of substrate specificity, both amylases could utilize all of the substrates tested with activity observed in the following order: amylopectin > potato starch > dextrin > glycogen > amylose. Statistical analysis indicated significant functional differences between the two allozymes in terms of specific activities, substrate specificities, and inhibitor sensitivities. AmyF had a significantly lower specific activity than did AmyS. The amylases responded differently to the substrate amylose, with AmyF better able to digest this substrate. AmyS was less sensitive than AmyF to α-amylase inhibitor from wheat.

Tissue-specific distribution of amylase in the mosquitofish (Gambusia affinis holbrooki).

Tissue-specific expression of alpha-amylase (E.C.3.2.1; α 1,4-glucan-4-glucano-hydrolase) was studied in the mosquitofish (Gambusia affinis holbrooki). Gut activity patterns, immunohistochemical distribution in the hepatopancreas and tissue-specific activities and amount of amylase protein were characterized. In the gut, activity was observed all along the gut and no variation in this pattern was observed in individuals from two natural populations. In the hepatopancreas, amylase is found in pancreatic tissue. Highest amylase activity was observed in gut tissue but highest amount of amylase protein was found in the hepatopancreas.

The amylase gene−enzyme system of chickens. I: Allozymic and activity variation

Amylase allozymic and activity variation was studied in three flocks of chickens (Gallus domesticus). Individuals from one flock were studied to assess the effects of sex, tissue, and genotype on amylase activity. Additionally, the allozymes were purified and their specific activities compared. Variation was observed within and among the flocks. Two alleles were found to be segregating in the flocks, one flock being polymorphic and the other two monomorphic. Mean amylase activities among the three flocks were significantly different. The relationship of this activity variation to regulatory variation is discussed. There were no significant effects of sex or genotype on amylase activity and, in most cases, no correlation between activities in the various tissues. However, in heterozygotes one of the alloamylases had much lower activity than the other.

Theamylase gene-enzyme system of fishes

SummaryAmylases from the mosquitofish (Gambusia affinis holbrooki, Pisces: Poeciliidae) and rat were purified and compared withDrosophila amylases in terms of structure and function. At the structural level, amino acid compositions of the three amylases were compared. At the functional level, amylase activities were compared on various substrates and in the presence of inhibitors.While the amylases from all three organisms had properties typical of alpha-amylases, both structural and functional differences were observed. Using resemblance coefficients of distance and similarity from numerical taxonomy, it was determined that the amylases from the rat andDrosophila were more similar to each other than either was to amylase from the mosquitofish, and that structural differences between the amylases did not reflect functional differences, i.e. there was no correlation between amylase structural and functional distances.

The amylase system of Drosophila. III: Biochemical comparison of amylases from D. pseudoobscura, D. persimilis, D. miranda and D. melanogaster

Abstract 1. 1. Six amylases from four species of Drosophila were compared in terms of specific activities, pH optima, substrate specificities, inhibitor sensitivities, responses to temperature, heat sensitivities, and cross-reactivity to amylase antiserum. 2. 2. The amylases studied were as follows: Amy1.00 and Amy0.84 from D. pseudoobscura; Amy1.00 and Amy1.09 from D. persimilis; Amy1.43 from D. miranda; and Amy6 from D. melanogaster. 3. 3. Both crude extracts and partially purified preparations were studied for some of the amylases. 4. 4. The amylases, in general, appeared quite similar to other amylases of Drosophila and were similar for most of their biochemical properties. Differences were observed, however, especially in regard to cross-reactivity to D. pseudoobscura Amy1.00 antiserum. 5. 5. These similarities and differences were discussed in relation to the ecological differences and the evolutionary (phylogenetic) relationships of the four species. 6. 6. Crude extract results corresponded, in general, to the results from the partially purified preparations. However, enough differences were observed to warrant caution when using crude extracts in attempting to characterize Drosophila amylases.