Genetical variation for enzyme activity in a population of Drosophila melanogaster. VI. Molecular variation in the control of alcohol dehydrogenase (ADH) activity

Four characters, ADH activity at 25 degrees, immunologically determined ADH protein level, total protein and body weight were measured upon 72 hour old adult female and male Drosophila melanogaster from 16 highly inbred lines, derived from the laboratory population, "Texas" (established 1966). The highest levels of ADH activity and ADH protein level were observed in the 2 lined homozygous for the AdhF allele. Amongst the 14 AdhS/S lines variation for ADH protein level was associated with genetical variation for ADH activity (r = 0.6). The genetical association between ADH activity or ADH protein level and either body weight or total protein in the 16 inbred lines was not statistically significant. A study of ADH activity, ADH protein and total protein in 8 lines representing all homozygous combinations of chromosomes I, II and III and derived from two inbred AdhS/S lines, chosen for their respective high and low ADH activities, showed that ADH activity was considerably modified by a post-translational event controlled from chromosome III. Total protein was controlled by different chromosomal effects from those controlling ADH activity. Michaelis constants for crude fly extracts of the two AdhF/F and the above two AdhS/S lines showed clear differences in affinity for isopropanol.     

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.     

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.     

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.     

The significance for breeding of linear regression analysis of genotype-environment interactions.

Methods of regression analysis of genotype-environment interaction are considered in relation to existing theory dealing with the relative efficiencies of selection for general or specific adaptation to the environment, and the choice of environments for assessment. The two alternative models is involving regression on to environmental effects (model 2) or genotypic effects (model 3) are equivalent when regression lines are concurrent, but are shown to be mutually exclusive when concurrence is absent...     

Genetic considerations in the effects of ethanol in mice. II. A trans-acting inducibility regulator(s) affecting alcohol dehydrogenase (ADH) activity

The inducibility of alcohol dehydrogenase (ADH) has been recognized in different systems including maize, Drosophila, and mice. Our earlier results showed strain-specific ADH responses to chronic ethanol administration relative to matched littermate controls in mice. For this study we used two strains which showed induction (BALB/c and S.W.) and two strains which showed repression (C57BL/6J and 129/ReJ) to produce three sets of F₁ hybrids and their reciprocals and one set (BALB/c×C57BL/6J) of recombinant inbred (RI) lines. The ADH properties of the resulting genotypes were again evaluated following 15% ethanol treatment in drinking water (2 weeks) in relation to their littermate matched controls in replicated trials. Our F₁ results suggest complete dominance for induction over repression at the phenotypic level, and the two repressed strains showed complementation. No significant differences were observed in the reciprocal F₁'s and all pairs of a given genotype-treatment combination yielded consistent results. The 1:1 segregation of RI lines suggests a single gene difference for ADH inducibility between BALB/c and C57BL/6J. These findings suggest the presence of a trans-acting inducibility regulator(s) for ADH which may or may not represent a single locus. Variability for such regulatory elements may provide an explanation for the commonly observed individual differences in natural populations for response to alcohol including alcohol metabolism.     

Ecological genetics of seed germination regulation in Bromus tectorum L.

The probability that a seed will germinate depends on factors associated with genotype, maturation environment, post-maturation history, and germination environment. In this study, we examined the interaction among these sets of factors for 18 inbred lines from six populations of Bromus tectorum L., a winter annual grass that is an important weed in the semi-arid western United States. Seeds of this species are at least conditionally dormant at dispersal and become germinable through dry-afterripening under summer conditions. Populations and inbred lines of B. tectorum possess contrasting dormancy patterns. Seeds of each inbred line were produced in a greenhouse under one of three levels of maturation water stress, then subjected to immediate incubation under five incubation regimes or to dry storage at 20°C for 4 weeks, 12 weeks, or 1 year. Dry-stored seeds were subsequently placed in incubation at 20/30°C. Narrow-sense heritability estimates based on parent-offspring regressions for germination percentage of recently harvested seeds at each incubation temperature were high (0.518–0.993). Germination percentage increased with increasing water stress overall, but there were strong interactions with inbred line and incubation temperature. Inbred lines whose seeds were non-dormant over the full range of incubation temperatures when produced at low maturation water stress showed reaction norms characterized by little or no change as a function of increasing stress. For inbred lines whose dormancy status varied with incubation temperature, incubation treatments where seeds exhibited either very low or very high levels of dormancy showed the least change in response to maturation water stress. Inbred lines also varied in their pattern of dormancy loss during storage at 20°C, but maturation water stress had only a minor effect on this pattern. For fully afterripened seeds (1 year in storage at 20°C), inbred line and maturation water stress effects were no longer evident, indicating that differences in genotype and maturation environment function mainly to regulate dormancy and dormancy loss in B. tectorum, rather than to mediate response patterns of non-dormant seeds.     

ADH enzyme activity and Adh gene expression in Drosophila melanogaster lines differentially selected for increased alcohol tolerance

In Drosophila melanogaster, alcohol dehydrogenase (ADH) activity is essential for ethanol tolerance, but its role may not be restricted to alcohol metabolism alone. Here we describe ADH activity and Adh expression level upon selection for increased alcohol tolerance in different life-stages of D. melanogaster lines with two distinct Adh genotypes: Adh(FF) and Adh(SS). We demonstrate a positive within genotype response for increased alcohol tolerance. Life-stage dependent selection was observed in larvae only. A slight constitutive increase in adult ADH activity for all selection regimes and genotypes was observed, that was not paralleled by Adh expression. Larval Adh expression showed a constitutive increase, that was not reflected in ADH activity. Upon exposure to environmental ethanol, sex, selection regime life stage and genotype appear to have differential effects. Increased ADH activity accompanies increased ethanol tolerance in D. melanogaster but this increase is not paralleled by expression of the Adh gene.     

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.     

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.     

Effects of genotype-environment interactions on genetic correlations

The objective of the work presented here was to investigate the influence of genotype-environment interaction on genetic correlations. In our theoretical models we have considered plant populations consisting of random samples of lines from chromosome-doubled haploids produced from F ₁ gametes, highly inbred SSD-lines, and clones of randomly breeding populations grown in two and multiple environments. The results of our theoretical considerations are that if genotype-environment interaction exists, great differences are expected to occur in the estimates of genetic correlation coefficients obtained in different environments. Based on the variance and covariance components for genotype-environment interaction we suggest a new type of correlation coefficient, called genotype-environment correlation, r _ge . Our theory has been applied to several series of experiments. Estimates are presented from two series, both of which demonstrate clearly the consequences of genotype-environment interaction on the genetic correlations.     

Performance of recombinant inbred lines in Brussels sprouts (Brassica oleracea var. gemmifera)

Summary Performance of a random array of recombinant inbred lines derived by single seed descent from five different source populations of Brussels sprouts (Brassica oleracea var. gemmifera) is presented. A total of 2,356 lines were tested in trials during 1985 and 1986. Three of the source populations were derived from double crosses between F₁ hybrids. These hybrids show a considerable heterotic advantage over their inbred parents for the most important agronomic traits. The recombinant inbred lines performed, on average, less well than the parental inbred material, indicating that additive x additive genie interactions may make a significant contribution to the performance of current inbred material. Nevertheless, the very large variation among the recombinant inbred lines permitted many lines to be identified which outperformed the best parental inbred for all traits. Two lines outperformed the reference F₁ hybrid, Gower, for an index that included marketable yield and quality. Consideration was also given to the dangers of misinterpreting phenotypically based proportions. Accordingly, response equations were used to ascertain the real genetic progress that was made. Advance seemed small when compared with the large heterotic effect, which is consistent with the segregation of a large number of loci. The distribution of the recombinant inbred lines was compared to predictions made from early generation trials. There was broad agreement but significant discrepancies existed which, it is suggested, may arise from the effects of genotype-environment interactions.     

Nonlatitudinal environmental correlations for alcohol dehydrogenase in Southern African populations ofDrosophila melanogaster

The relation between alcohol dehydrogenase (ADH) allozyme frequencies and several environmental variables, as well as latitude and longitude, was examined in 17 natural populations of Drosophila melanogaster in Namibia, South Africa, and Zimbabwe. Unlike several other studies, we found no latitudinal allele-frequency clines and no significant correlation with temperature. We found that the frequency of the most common allele (AdhS) was positively correlated with an increase in rainfall. We suggest that substrate-induced toxicity may explain the geographic variability in our results.     

The Influence of Donor Plant Genotype and Environment on Production of Multicellular Microspores in Cultured Anthers of Brassica napus ssp. oleifera

Studies of the effects of genotype and pre-flowering environmentalconditions on the production of multicellular microspores wereundertaken th four highly inbred lines of Brassica napus sap.oleifera. These lines were first grown in shaded and unshadedenvironments at 20/15°C arid unshaded at 30/25°C ina daylight phytotron. Buds were harvested from half the plantswhen first visible in the rosette and later from the remainingplants at the time when the first flower opened. The frequencyof microspores at a specific stage of development varied widelywithin a relatively narrow range of bud lengths. Uninucleatemicrospores were not detected in anthers from buds less than1·5 m or greater than 3·0mm long, but were generallypresent in frequencies of greater than 50 per cent in anthersfrom buds which were between 2·0 and 2·5 mm inlength. However, the bud length at which the highest frequencyof uninucleate microspores was detected varied significantlybetween genotypes and between the environments in which theywere grown. Examination of the remaining anthers from each budafter a period in culture revealed that the proportion of microsporesdeveloping into multicellular units varied greatly with budlength, an increase in frequency of multicellular microsporesbeing associated with an increase in the frequency of uninucleatemicrospores in the uncultured anther. Genotypes differed, however,in respect of the relationship between uninucleate microsporefrequency and production of multicellular units. Although thefrequency of multicellular units was as high as 57 percent,further development was limited and the number of embryoldsformed was low in all cases (<10 per cent). The frequency of multicellular units in pollen samples frombuds of a length in which uninucleate microspore frequency washigh varied significantly with genotype, temperature and lightconditions under which donor plants were grown, and the stageof inflorescence development at which buds were removed. Underconditions most conducive to multicellular unit formation (20/15°C,unshaded), the maximum frequency of multicellular units foreach genotype in buds from young inflorescences ranged from11·5 to 56·5 per cent. Shading or exposure tothe higher temperature was associated with a marked reductionin production of multicellular units. Higher frequencies ofmulticellular units were generally detected in microspore samplesfrom younger inflorescences irrespective of genotype or environment. Two of the four inbred lines were selected for a second experimentin which responses to vernalization and photoperiod durationwere monitored. There was a significant reduction in the numberof leaf nodes formed prior to floral initiation in both genotypesfollowing exposure to vernalization and/or a longer photoperiod,the response to photoperiod being more pronounced. Exposureto 4 weeks vernalization was accompanied by a significant increasein the frequency of multicellular units in both genotypes, thefrequency being double that in unvernalized plants under thelonger photoperiod. By contrast, genotypes differed sharplyin their response to photoperiod. In TB 20, the frequency ofmulticellular units was unaffected by an increase in day lengthirrespective of whether seed had been vernalized or not. Onthe other hand, in TB 42 the frequency of multicellular unitswas substantially greater in the 24 h day than in the 12 h day,being 27·3 per cent vs 13·0 per cent in the caseof unvernalized plants and 66·7 per cent vs 18·2per cent in the case of vernalized plants. Brassica napus, anther culture, pollen embryogenesis, genotype-environment interaction     

An epidemiologic approach to ecogenetics.

Although "ecogenetics" seeks to examine genetically mediated differences in susceptibility to environmental agents, researchers often examine the relation between genetic markers and disease without regard to environmental determinants. By using epidemiologic definitions of genotype-environment interaction, it can be shown that the relative risk of disease for the genetic marker is a function of the frequency of exposure to the environmental agent, the strength of interaction between the genotype and the agent, and the specificity of the environmental effect vis-à-vis the genotype. Using examples from the literature, we illustrate under six patterns of genotype-environment interaction that the relative risk associated with the marker can fluctuate markedly. However, with infrequent exposures, the relative risk is close to unity (implying no genetic effect) even in the face of strong genotype-environment interaction. Alternatively, elevated relative risks imply a frequent environmental exposure or a strong pattern of interaction. We suggest that genetic marker-disease associations be evaluated within the context of an epidemiologic study design that considers specific environmental determinants of risk.     

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.     

Effects of inbreeding on economic traits of channel catfish

Summary Inbred channel catfish (Ictalurus punctatus) were produced from two generations of full-sib matings to study the effect of inbreeding on reproduction, growth and survival. A randomly mated control line was propagated from the same base population to be used for the evaluation of the inbred fish. First generation inbred (I1) and control (C1) lines comprised five full-sib families each. Second generation inbred (I2) and control (C2) lines were produced by mating each male catfish from the I1 or C1 line to two females in sequence, one from the I1 and one from the C1 line. The design also produced two reciprocal outcross lines to be compared to their contemporary inbred and control lines. The coefficient of inbreeding for the inbred line increased from 0.25 in generation 1 to 0.375 in generation 2. The inbreeding coefficient was zero for all other lines. The resulting fish were performance tested in two locations, Tifton, Georgia and Auburn, Alabama and no genotype-environment interactions occurred. Results indicated that one generation of inbreeding increased number of days required for eggs to hatch by 21%, but did not significantly influence spawn weight or hatchability score. However, inbred females produced more eggs/kg body weight than control females. Two generations of full-sib mating in Georgia did not depress weight when expressed as a deviation to random controls but was depressed 13–16% when expressed as a deviation to half-sib out-crosses. Second generation inbreds produced in Alabama exhibited a 19% depression for growth rate when compared to either random or half-sib outcross controls. Survival rates at various age intervals was not decreased by inbreeding. The amount of inbreeding depression varied among families and between sexes.This study was supported by State and Hatch Funds allocated to the Georgia and Alabama Agricultural Experiment Stations