Genetical variation for enzyme activity in a population of Drosophila melanogaster. VII. Genotype-environment interaction for alcohol dehydrogenase (ADH) activity

Genotype-environment interaction was detected for ADH activity amongst a set of 18 highly inbred lines of Drosophila melanogaster which had been extracted from the laboratory population, "Texas". The genotype-environment interaction for ADH activity was not wholly associated with genotype-environment interaction for body weight or total protein level. Detailed analyses of the responses of the individual inbred lines in ADH activity in relation to the environmental index, ej and following the procedure of Jinks and Pooni (1979), showed substantial diversity in the form of response. Lines homozygous for the AdhF allele were more environmentally sensitive than AdhS/S lines. Amongst the 16 AdhS/S lines, models of linear, quadratic or two intersecting-straight-lines were used to illustrate the varied responses of genotypes to the environment. The heterogeneity in the response characteristics of the inbred lines was attributed to variations in the conditions of culture media normally present within populations and laboratories. Moreover the non-linear responses shown by some lines to the environment are consistent with a model of genotype-environment interaction for ADH activity mediated by varied genotype-specific sensitivities to different environmental factors.     

Genetical variation for enzyme activity in a population of Drosophila melanogaster. IV. Analysis of alcohol dehydrogenase activity in chromosome substitution lines.

Chromosome substitution lines derived from two inbred strains of Drosophila melanogaster homozygous for the AdhS allele of alcohol dehydrogenase but differing significantly in ADH activity have been analysed. Variation in activity can be attributed to all three major chromosomes. The effect of the second chromosome, where the ADH structural gene is located, can be modified significantly by the genotype of both the first and the third chromosomes. The most substantial single effect results from homozygous differences between the third chromosomes. In contrast, differences between the X chromosomes are revealed only when the second or second and third chromosomes are heterozygous.     

Effects on ADH activity and distribution, following selection for tolerance to ethanol in Drosophila melanogaster.

Strains of Drosophila melanogaster homozygous for either the AdhF or the AdhS allele were kept on food supplemented with ethanol for 20 generations. These strains (FE and SE) were tested for tolerance to ethanol and compared with control strains (FN and SN). The E strains showed increased tolerance to ethanol both in the adult and in the juvenile life stages. In adults the increase in tolerance was not accompanied by an increase in overall ADH activity. However, there were changes in the distribution of ADH over the body parts. Flies of the FE strain possessed significantly more ADH in the abdomen, compared with FN. Another set of FN and SN populations were started both on standard food and on ethanol food with reduced yeast concentrations. After 9 months ADH activities were determined in flies from these populations which had been placed on three different media: the food the populations had been kept on, regular food and regular food supplemented with ethanol. The phenotypic effects of yeast reduction on ADH activity were considerably, but longterm genetic effects were limited.     

The partial characterization of alcohol dehydrogenase null alleles from natural populations ofDrosophila melanogaster

Twenty-three alcohol dehydrogenase (ADH) putative null alleles extracted from four Tasmanian (Australia) populations of Drosophila melanogaster produce no ADH activity and are unable to form active heterodimers with either AdhF or AdhS. Twelve of these nulls were tested by enzyme-linked immunosorbent assay (ELISA) and did not produce any ADH cross-reacting material (CRM). The null homozygotes had similar, but slightly lower, mortalities on ethanol-supplemented media compared to an artificially induced null allele. Heterozygotes between the null alleles and standard AdhF and AdhS alleles had intermediate ADH activity and CRM levels.     

Biochemical differences between alcohol dehydrogenases of Drosophila melanogaster.

This paper describes substrate specificities, developmental changes in activity, pH profiles, and heat stabilities of isozymes produced by four Adh genotypes in D. melanogaster. No differences are found in the substrate specificities of isozymes from the different genotypes but studies of the other three properties reveal significant differences between the isozymes. Thus relatively low activities are found among extracts of AdhF Adhn2 larvae and among extracts of AdhF AdhF adults aged 44 days. Also AdhF AdhS and AdhS AdhS extracts have relatively high activities at pH 6-5, and AdhF Adhn2 extracts have relatively low activities at pH values above 10-0. Finally, extracts of AdhF AdhF and AdhF AdhS are more stable at 40 degrees C than are those of AdhS AdhS and AdhF Adhn2.     

Selection for ethanol tolerance in two populations of Drosophila melanogaster segregating alcohol dehydrogenase allozymes.

Selection for ethanol tolerance was equally successful in two populations of D. melanogaster in both of which the frequency of AdhF was 0.5 at the start of the experiment. Increased tolerance to ethanol was not invariably associated with increased frequencies of AdhF. In one population alcohol dehydrogenase (ADH) activity was significantly higher in three of the four selected sublines compared with their controls but there was no difference in activity between the selected and control sublines in the second population. The level of ADH activity in the control and selected lines was significantly correlated with the frequency of AdhF, but not with ethanol tolerance. These results show that adaptation to environmental alcohols in populations of D. melanogaster can be independent of the ADH system.     

Tolerance to 1-pentene-3-ol and to 1-pentene-3-one in relation to alcohol dehydrogenase (ADH) and aldo keto reductase (AKR) activities inDrosophila melanogaster

The detoxification of 1-pentene-3-ol (pentenol) and 1-pentene-3-one (pentenone) by Drosophila melanogaster adult flies has been studied in two homozygous lines for the AdhF and AdhS alleles (LRC lines), in their respective lines selected for tolerance to ethanol (LRSe lines) and in a homozygous strain for the Adhn4 null allele. For each line, the genotype and sex LDs50 of both compounds were estimated. Then, in order to explain the differences in LD50, both alcohol dehydrogenase (ADH) and aldo keto reductase (AKR) activities were assayed. In addition, the effects of pentenone on AKR activity were also studied. Our results show that ADH-positive flies exhibit a much higher sensitivity to pentenol than ADH-null flies. However, both ADH-positive and ADH-null flies show a similar tolerance to pentenone. Our results show that flies selected for improving tolerance to ethanol also have increased tolerance to pentenol (FF and SS flies) and pentenone (SS flies). However, this improved ability to tolerate pentenol and/or pentenone cannot be explained by changes in ADH or AKR activities. On the other hand, we have observed a beneficial effect of pentenol, but not of pentenone, in n4 flies. We also show that AKR activity is not modified by the administration of pentenone. These results suggest that, in the absence of ADH activity, pentenol may be transformed into a compound that is less toxic than pentenone and that pentenone itself might also be transformed into a less toxic compound.     

Hepatic aldehyde dehydrogenase activity in Peromyscus genetically deficient in alcohol dehydrogenase

1. Hepatic aldehyde dehydrogenase (ALDH) activity was measured in two strains of deer-mouse, Peromyscus maniculatus. 2. There is no difference in the subcellular distribution of ALDH activity in the two strains. Animals of AdhN/AdhN genotype, lacking liver alcohol dehydrogenase (ADH), had 90% of total ALDH activity in the mitochondrial fraction compared to 94% for the AdhF/AdhF animals with normal ADH activity. Almost all of the remaining ALDH activity was in the hepatic cytosol with less than 1% in the microsomal fraction. 3. By contrast, in mice (Mus musculus) 43% of total hepatic ALDH activity was found in the cytosolic fraction and 55% in the mitochondrial. 4. It was concluded that the subcellular distribution of hepatic ALDH activity in Peromyscus does not vary with the presence or absence of ADH and that this ALDH distribution is not similar to that reported for other rodents.     

Physiological Significance of the Alcohol Dehydrogenase Polymorphism in Larvae of Drosophila

This study deals with biochemical and metabolic-physiological aspects of the relationship between variation in in vivo alcohol dehydrogenase activity and fitness in larvae homozygous for the alleles Adh71k, AdhF, AdhS, of Drosophila melanogaster, and for the common Adh allele of Drosophila simulans. The Adh genotypes differ in the maximum oxidation rates of propan-2-ol into acetone in vivo. There are smaller differences between the Adh genotypes in rates of ethanol elimination. Rates of accumulation of ethanol in vivo are negatively associated with larval-to-adult survival of the Adh genotypes. The rank order of the maximum rates of the ADHs in elimination of propan-2-ol, as well as ethanol, is ADH-71k greater than ADH-F greater than ADH-S greater than simulans-ADH. The ratio of this maximum rate to ADH quantity reveals the rank order of ADH-S greater than ADH-F greater than ADH-71k greater than simulans-ADH, suggesting a compensation for allozymic efficiency by the ADH quantity in D. melanogaster.Our findings show that natural selection may act on the Adh polymorphism in larvae via differences in rates of alcohol metabolism.     

Gene-enzyme system of alcohol dehydrogenase and adaptation of Drosophila to increased temperature]

Properties and allelic ADH (alcohol dehydrogenase) control in Drosophila melanogaster were studied upon flies' cultivation under conditions of hypotherm of different intensity and duration. Lines homozygotic for F allele (vg) and S allele (cn) of the Adh gene as well as genetically enriched experimental cn' and vg' populations containing a small number of AdhF/AdhS heterozygotes at the initial stage were used. It was found out that physiological adaptation of the species to momentary influence of elevated temperature is accompanied by modification of physical properties of ADH-F according to ADH-S. Constant influence (during the life span of 25 generations) of elevated temperature on the population changes its genetic structure, due to selective advantages of the S allele of Adh under these conditions.     

Genotype-environmental interaction in the activity and preening of Drosophila melanogaster

Summary Techniques, recently developed to analyze genotype-environmental interaction in plants, are used to study the behaviour of two inbred lines of Drosophila melanogaster and their F ₁s. The locomotor activity and preening of the hybrids altered far less with age and between two different test conditions than did the behaviour of their homozygous parents. In one apparatus, age affected the additive genetical component leading to heterosis for high activity and, in the other condition, a maternal effect on activity was age-dependent. Preening varied far less with age but, like activity, showed dominance for low inter-individual variability and differences between replicates, due entirely to the unstable performance of one inbred line.In the light of additional evidence on larval-adult survival and on adult viability, it is suggested that a negative correlation exists between viability and responsiveness to many types of environmental variation. Therefore information may be gained by analyzing inter-individual variability, instead of the customary practice of rescaling data to make the variances homogeneous.     

Genetic considerations in the effects of ethanol in mice. I. Genotype-dependent alterations in alcohol dehydrogenase activity

Most genetic studies on individual and racial differences in sensitivity to alcohol intoxication have concentrated on genetic variations associated with structural genes for the enzymes involved in alcohol metabolism, including alcohol dehydrogenase (ADH; E.C. 1.1.1.1). We studied the ethanol-induced regulation of ADH following chronic administration of ethanol in mice. Newly weaned males from six inbred strains (BALB/c, C3H/HeSnJ, C3H/S, C57BL/6J, S.W., and 129/ReJ) were subjected to ethanol administration. Alterations in the level of liver ADH activity, relative to matched littermate controls, were evaluated. The change in ADH activity was found to be strain (genotype) specific, which may explain the contradictory results in the literature. Strains which showed induction of ADH activity, in general, reflected a strain-specific time-dependent profile. Strains which showed repression, however, were independent in the degree of repression to the duration of ethanol exposure. Such variable, ethanol-induced regulatory responses (induction/repression) in ADH activity of different genotypes may account for individual and population variations in response to alcohol. Additional work, however, is needed to establish the molecular bases of ADH inducibility and its specific role in relative susceptibility to alcohols.     

Comparison of alcohol dehydrogenase expression in Drosophila melanogaster and D. simulans

Alcohol dehydrogenase (ADH) gene expression was analyzed in Drosophila melanogaster and its sibling species D. simulans. The levels of ADH activity, ADH-cross-reacting material (CRM), and ADH-mRNA were analyzed for several strains of each species, which derive from diverse geographic locations around the world. There is considerable quantitative variation in ADH activity, CRM level, and RNA level among strains within species at all developmental stages. However, the only consistent differences between the two species are in pupal RNA level and in late-adult activity and CRM level. Late-adult melanogaster flies that are homozygous for the Slow allozyme have approximately twice the level of ADH activity and CRM as do simulans flies. The regression of activity on CRM over strains is highly significant and essentially the same for each species, which means that most, if not all, of the activity difference between the species is due to a difference in concentration of the ADH protein. In contrast, there is no significant regression of CRM level on mRNA level in adults of either species; nor is there a significant difference in RNA level between species. Therefore, the difference in ADH protein concentration is not due to RNA template availability. Thus, the interspecific difference in ADH level in adults must be due either to a difference in the rate of translation of the two RNAs or to a difference in protein stability.     

Peromyscus alcohol dehydrogenase: Lack of cross-reacting material in enzyme-negative animals

Two forms of alcohol dehydrogenase (ADH), coded by allelic genes, have been purified to homogeneity from Peromyscus. Monospecific antisera to the purified enzymes have been raised in rabbits. These antisera fail to detect cross-reacting material in the liver of ADH-negative animals on Ouchterlony plates. Immuno-titration of anti-ADH antiserum with ADH in liver extracts from AdhS/AdhS and AdhS/AdhN animals results in identical equivalence points, again suggesting the absence of cross-reacting material coded by the AdhN allele. Over a wide range of anti-ADH antiserum dilutions, radiolabeled protein was not immunoprecipitable from liver extracts of AdhN/AdhN animals. These immunochemical tests, in conjunction with previous studies, suggest that the AdhN allele in Peromyscus does not produce inactive polypeptide in normal levels that bears immunological determinants similar to those of the fast and slow ADH isozymes.     

Efficiency of selection, as measured by single nucleotide polymorphism variation, is dependent on inbreeding rate in Drosophila melanogaster

It is often hypothesized that slow inbreeding causes less inbreeding depression than fast inbreeding at the same absolute level of inbreeding. Possible explanations for this phenomenon include the more efficient purging of deleterious alleles and more efficient selection for heterozygote individuals during slow, when compared with fast, inbreeding. We studied the impact of inbreeding rate on the loss of heterozygosity and on morphological traits in Drosophila melanogaster. We analysed five noninbred control lines, 10 fast inbred lines and 10 slow inbred lines; the inbred lines all had an expected inbreeding coefficient of approximately 0.25. Forty single nucleotide polymorphisms in DNA coding regions were genotyped, and we measured the size and shape of wings and counted the number of sternopleural bristles on the genotyped individuals. We found a significantly higher level of genetic variation in the slow inbred lines than in the fast inbred lines. This higher genetic variation was resulting from a large contribution from a few loci and a smaller effect from several loci. We attributed the increased heterozygosity in the slow inbred lines to the favouring of heterozygous individuals over homozygous individuals by natural selection, either by associative over‐dominance or balancing selection, or a combination of both. Furthermore, we found a significant polynomial correlation between genetic variance and wing size and shape in the fast inbred lines. This was caused by a greater number of homozygous individuals among the fast inbred lines with small, narrow wings, which indicated inbreeding depression. Our results demonstrated that the same amount of inbreeding can have different effects on genetic variance depending on the inbreeding rate, with slow inbreeding leading to higher genetic variance than fast inbreeding. These results increase our understanding of the genetic basis of the common observation that slow inbred lines express less inbreeding depression than fast inbred lines. In addition, this has more general implications for the importance of selection in maintaining genetic variation.     

Genetic variation in the pleiotropic association between physical activity and body weight in mice

Background

A sedentary lifestyle is often assumed to lead to increases in body weight and potentially obesity and related diseases but in fact little is known about the genetic association between physical activity and body weight. We tested for such an association between body weight and the distance, duration, and speed voluntarily run by 310 mice from the F₂ generation produced from an intercross of two inbred lines that differed dramatically in their physical activity levels.

Methods

We used a conventional interval mapping approach with SNP markers to search for QTLs that affected both body weight and activity traits. We also conducted a genome scan to search for relationship QTLs (relQTLs), or chromosomal regions that affected an activity trait variably depending on the phenotypic value of body weight.

Results

We uncovered seven quantitative trait loci (QTLs) affecting body weight, but only one co-localized with another QTL previously found for activity traits. We discovered 19 relQTLs that provided evidence for a genetic (pleiotropic) association of physical activity and body weight. The three genotypes at each of these loci typically exhibited a combination of negative, zero, and positive regressions of the activity traits on body weight, the net effect of which was to produce overall independence of body weight from physical activity. We also demonstrated that the relQTLs produced these varying associations through differential epistatic interactions with a number of other epistatic QTLs throughout the genome.

Conclusion

It was concluded that individuals with specific combinations of genotypes at the relQTLs and epiQTLs might account for some of the variation typically seen in plots of the association of physical activity with body weight.     

Genetic variation in the expression of ADH in Drosophila melanogaster

Several chromosomes derived from natural populations have been identified that affect the expression of alcohol dehydrogenase (ADH). Second chromosomes, which also carry the structural gene Adh, show a great deal of polymorphism of genetic elements that determine how much enzyme protein accumulates. The level of enzyme was measured in third instar larvae, 6-to-8-day-old males and in larval fat bodies and alimentary canals. In general, activities in the different organs and stages are highly correlated with one another. One line was found, however, in which the ADH level in the fat body is more than twice the level one would expect on the basis of the activity in alimentary canal. We have also found evidence of third-chromosome elements that affect the level of ADH.     

Somaclonal variation in some agronomic and quality characters in wheat

Summary A total of 256 selected lines derived from tissue culture of three hexaploid wheat cultivars were grown in a replicated hill plot experiment to examine somaclonal variation in agronomic characters. The lines were derived by single plant selection for various characters from a total of 100 regenerants, and were either SC₃ or SC₄ generation in the test. Significant variation was found in all the characters measured: height, grain number per spike, kernel weight, yield, total dry weight and harvest index. In most cases, variation could be identified which was both less than and greater than the parental controls. However, there was also an apparent effect of the parent cultivar on the total amount and direction of the variation. For two cultivars, lines could be traced back through the culture phase to individual explant embryos. Many of the original embryos contributed significant variation, and most characters showed significant variation arising from more than one embryo. In the following year, 32 lines selected from the hill plot experiment were grown in larger replicated plots and yield, harvest index and a number of grain and baking quality characters were measured. No lines selected for high yield or harvest index maintained significant improvements over their parental controls. However, significant variation was displayed for many of the quality characters examined. Significant increases in kernel weight, hardness and protein content, and a significant reduction in yellow pigmentation represented potentially useful improvements. Only unfavourable variation was seen in flour yield and in mixograph height, time and breakdown.     

Structure of variation in enzyme activity in natural Drosophila melanogaster populations

Enzyme activity variation was assessed in several isofemale lines originating from two Hungarian Drosophila melanogaster populations. Samples from each population were taken from from two villages; 8-9 isofemale lines were established from each village. The activities of ADH, alphaGPDH, IDH and 6PGDH were determined in the adults (in the F1 generation) and in the larvae (in the F3 generation) as well. Enzyme activities were measured on starch gel after electrophoresis. The activity of the enzyme was detected in a single individual and it was also possible to determine its genotype. The results showed that most of the variation occurred within sites for all four enzymes. This within site variation was more or less equally partitioned into within and between isofemale line (family) components. A smaller portion of variation was attributable to the differences between the populations. Nevertheless, adult alphaGPDH, and larval IDH and 6PGDH activities exhibited significant differences between the two populations. Variation in larval activities of all enzymes was higher than that of the adults, but 6PGDH had considerably higher variation in the adults. The greater variation in larval activities probably reflected the greater environmental variation in the microhabitat of the larvae compared to that of the adults. Larval activities of the investigated enzymes showed much stronger correlation than adult activities. The correlation pattern in the adults differed greatly between the two populations.