THE ASSOCIATION BETWEEN THE POLYMORPHISMS AT THE Adh AND αGpdh LOCI AND THE In(2L)t INVERSION IN DROSOPHILA MELANOGASTER IN RELATION TO TEMPERATURE

Substantial allele-frequency changes were observed at the Adh and αGpdh loci in a seminatural population of Drosophila melanogaster kept in a tropical greenhouse during 1972–1985. Further analysis of the changes at the Adh and αGpdh loci showed that linkage disequilibrium between these loci occurred for a prolonged period due to the presence of In(2L)t, a long inversion on the left arm of the second chromosome. We observed increases in the frequencies of In(2L)t and of short inversions on the left arm of the second chromosome in subpopulations kept at 29.5°C or 33°C. These inversion-frequency increases were accompanied by an increase in Adh^s and a decrease in αGpdh^s frequency. In populations kept at 20°C and 25°C, inversion frequencies decreased, while αGpdh^s allele frequencies decreased at 25°C and increased at 20°C. At 33°C, egg-to-adult survival of individuals possessing In(2L)t, either in the homokaryotypic or the heterokaryotypic state, was higher than that of the other karyotypes of identical allozyme constitution (i.e., Adh^s αGpdh^F). Thus it seems that In(2L)t has a selective advantage at high temperature. We argue that the observed changes in allele frequencies at the Adh and αGpdh loci are, in part, due to genic selection and are not merely the result of selection acting on the chromosome rearrangements and hitchhiking of the allozymes. The results are discussed with respect to the latitudinal clines found for In(2L)t, Adh, and αGpdh.     

Association of Chromosome and Enzyme Polymorphisms in Natural and Cage Populations of DROSOPHILA MELANOGASTER

The frequencies of a polymorphic inversion, In(2L)t, and of Adh and αGpdh alleles were analyzed in three natural populations of Drosophila melanogaster from Japan. Significant positive correlations between the frequencies of In(2L)t and Adh^S or αGpdh^F were detected due to tight linkage. An analysis of correlation with latitude showed that the negative cline of Adh^S frequency could be explained entirely by its linkage with In(2L)t; the frequency of Adh^S on the standard chromosome did not show a latitudinal cline. To the contrary, the cline of αGpdh^F frequency itself was positive, and its linkage with In(2L)t makes the positive cline unclear. These results suggest that the two allozymes themselves respond to latitudinal natural selection in different ways. When these populations were transferred to laboratory cages and maintained for a long time, they lost the chromosomal polymorphism but retained stable enzyme polymorphisms, although allele frequencies in the cage were not the same as in nature. The frequencies of Adh and αGpdh alleles were close to those in earlier cage populations of the same geographical origin.     

Chromosome inversion polymorphisms in Drosophila melanogaster III. Gametic disequilibria and the contributions of inversion clines to the Adh and Gpdh clines in Australasia

Gametes from each of 19 Australasian collections of Drosophila melanogaster were characterised for polymorphic Adh and Gpdh allozyme loci and the two chromosome-2 common cosmopolitan inversions In(2L)t and In(2R)NS.-No consistent gametic disequilibria for the loosely linked combinations of Adh-Gpdh and Adh-In(2R)NS were found over the 19 collections. However, strong and consistent gametic disequilibria were found for the tightly linked combinations of Adh-In(2L)t and Gpdh-In(2L)t. The results for each of these four combinations agree with those previously reported for Northern-Hemisphere collections. -Notwithstanding its parallel latitudinal cline and consistent gametic disequilibrium with Adh, In(2L)t was found to account for only a small fraction of the Adh latitudinal cline in Australasia. This is consistent with the previous finding that the North American Adh latitudinal cline also exists largely independently of In(2L)t. However, In(2L)t accounts for all of the Australasian Gpdh latitudinal cline and so contrasts the finding from North American collections where the corresponding Gpdh cline exists largely independently of In(2L)t.     

COMPETITION AND GENOTYPE-BY-ENVIRONMENT INTERACTION IN NATURAL BREEDING SUBSTRATES OF DROSOPHILA

Although empirical studies frequently suggest that genotype-by-environment (G X E) interaction can maintain genetic variation, very few data are available to test for the specific conditions necessary for the existence of a protected polymorphism (i.e., the property of persistence of an allele even when initially rare). Drosophila species live in patchy environments and their local population structure may be characterized to some extent by Levene's migration pattern, namely by a single pool of individuals that presumably mate at random and breed on discrete and ephemeral resources. We present here a field experiment that links Drosophila ecology and population genetics, which used the alcohol dehydrogenase (Adh) and α-glycerophosphate dehydrogenase (αGpdh) polymorphic loci in D. melanogaster flies raised from Opuntia ficus-indica fruits (prickly pears). The results show that there is density-dependent mortality in those fruits with a relatively high number of larvae (i.e., selection is “soft”) and suggest that there is differential viability for αGpdh genotypes. Additionally, a pattern of G X E interaction for fitness values, which is fully compatible with the theoretical conditions required for the existence of a protected polymorphism, was found after weighting the fitness estimates by the relative contribution that each fruit makes to the total adult population. The strong association between Adh^S and αGpdh^F alleles suggests that the occurrence of the common cosmopolitan inversion In(2L)t in the population might be responsible for the negative frequency-dependent selection predicted by Levene's model when genetic variation persists in heterogeneous environments.     

Allozyme variation in Greek wild populations of Drosophila melanogaster and D. simulans along a North-South gradient

Allozyme frequency data from five Greek wild sympatric populations of Drosophila melanogaster and D. simulans along a North-South gradient were analyzed for genotype-environment relationships. The regression coefficient of genetic distance on geographic distance indicates that there is a significant relationship between these parameters for D. melanogaster only. Highly significant differences in specific alleles at certain loci were found between the various local populations studied. The changes in Gpdh ^F of D. melanogaster and Est-6 ^F of D. simulans exhibited clinal patterns in allele frequencies. In addition, analysis of D. melanogaster Gpdh ^Fand Adh ^F allele frequencies shows that the Greek data do not have regression coefficients (regressing allele frequency on degrees North of latitude) of the same sign as East-and West-Coast United-States populations. These contradictory data are discussed in relation to what is known about the maintenance of the Adh and Gpdh polymorphisms.     

Selection against homozygotes in the ADH polymorphic system of Drosophila melanogaster associated with the lethal factor l(2) Stm

In the natural populations ⁺Tüb, ⁺Prov, and ⁺Rov, similar Adh ^F allele frequencies occur (q _F=0.11, 0.18, and 0.08, respectively). However, there is a discrepancy in that the Adh ^F allele in ⁺Tüb is closely linked to the lethal factor 1(2)Stm, which reduces relative fitness of the F phenotype to zero. In spite of this, polymorphism is maintained also in ⁺Tüb, because the heterozygotes are superior to the homozygous S type (relative fitness=0.88). Under laboratory culture conditions, in ⁺Tüb the relative fitness of the S genotype further decreases to 0.6. After outcrossing the lethal factor, relative fitnesses for S, FS, and F become 0.6, 1, and 0.48, respectively, implying that fitness for S remains the same. Relative values for S, FS, and F in ⁺Prov, not affected by the lethal factor, are calculated by the maximum average fitness method to be 1, 1.2, and 0.2 under the assumption that heterozygous FS are similarly superior to S as in the natural ⁺Tüb population and all allele frequencies found are stable equilibrium values.     

Alcohol dehydrogenase allozymes in the safflower genus Carthamus L.

The ADH allozyme pattern was tested in seeds of 1553 varieties of the world collection of safflower (Carthamus tinctorius L.) and 36 collections belonging to 14 wild species of the genus Carthamus L. with different chromosome numbers (n=10, 11, 12, 22, and 32). Two genes, Adh ₁ and Adh ₂, have been identified. The Adh ₁ locus controls the allozyme bands found in the faster-moving anodal zone, and the Adh ₂ gene controls the cathodal band. A third group of bands which migrates slowly toward the anode and stains weakly is probably interaction products of the two genes. Two codominant alleles Adh ₁ ^S and Adh ₁ ^F , specifying allozymes with different migration rates in the fast-moving anodal zone, were found in cultivated safflower. The frequency of the Adh ₁ ^F allele was very low. A third homologous allele, Adh ₁ ^T , was present only in the polyploid wild species. The Adh ₂ was stable, without any variation in migration rate. In addition to the variation in migration rates, there was also variation in activity levels of the products of both the Adh ₁ and Adh ₂ genes. The contribution of this study to our understanding of the origin of the polyploid species C. lanatus, C. baeticus, and C. turkestanicus is discussed.This research has been financed in part by a grant made to A. Ashri by the U.S. Department of Agriculture, Agricultural Research Service, authorized by P.L. 480, Project No. A10-CR-18, Grant No. FG-Is-234.Based in part on a thesis submitted by M. P. to the Faculty of Agriculture, The Hebrew University, Rehovot, in partial fulfillment of the requirements for the M.Sc. degree.Graduate Student, Faculty of Agriculture.     

Association and the Adh locus with a lethal factor (l(2) Stm) in Drosophila melanogaster

Keeping Drosophila cultures at 28 C results in elimination of all minor multiple ADH bands, thought to be due to conformational change. Thus in diploid and triploid adults heterozygous for the Adh ^F and Adh ^Salleles, relative staining intensities are found for the three bands which were in conformity with the assumption that both alleles are equally expressed. Among all polymorphic strains derived from natural Central European and Mediterranean populations, the strain ⁺Tüb is unique in that its Adh ^Fallele is closely linked to a new recessive lethal factor, named 1(2)Stm. All Adh ^F 1/Adh^F 1 pupae are unable to emerge, and die. The lethal effect is obvious 50 hr earlier by retarded eye, bristle, and body wall pigmentation. Although all pupae of the phenotype F die, Adh ^F allele frequency scarcely seems to be lowered in this natural population.     

Analysis of microdifferentiation in a Spanish cellar population of Drosophila melanogaster

Variation in Adh and Gpdh-1 gene frequencies has been used to check for microdifferentiation in Spanish samples of Drosophila melanogaster inside and outside a wine cellar. Flies were collected after vintage and after overwintering respectively; within each period samples were taken on up to five consecutive days each month. Variation of gene frequencies of Adh and Gpdh-1 can be considered random when samples collected each month are taken into account. When mean monthly frequencies are considered, Gpdh-1 does not show any significant variation all over the year; yet, variation of the frequency of Adh ^S shows a cyclical pattern, its frequency being maximum at the end of the summer and minimum after overwintering. Due to the parallel change of the frequency of the inversion In(2L)t and the Adh ^S allele, no decision can be made whether the Adh locus itself or the inversion are responsible for the changes.     

Association of allozyme heterozygosity and sternopleural chaetae number inDrosophila melanogaster

Variation at two polymorphic allozyme loci (Adh and Gpdh) has been assayed in relation to sternopleural chaetae number in flies derived from a cage population ofD. melanogaster known to be under stabilising selection with respect to chaetae number. Central phenotypes were significantly more heterozygous at these loci than more extreme phenotypes. Analysis of the base popylation and of divergent directional selection lines derived from it suggested that alleles atAdh and Gpdh have a pleiotropic effect on chaetae number or are in strong linkage disequilibrium with chaetae factors. In addition, homozygotes atAdh and Gpdh were observed to have a higher variance in chaetae number than heterozygotes. It is concluded that additive and homeostatic models of gene action might both partly explain the higher heterozygosity of central phenotypes.     

Enzymatic responses of Drosophila melanogaster to long- and short-term exposures to ethanol

The effects of environmental ethanol on larva-to-pupa survival and on the activities of four enzymes were investigated in three Drosophila melanogaster strains. The strains had different allelic combinations at the Odh and Aldox loci on their third chromosomes, but they all carried the Adh ^S -Gpdh ^F allelic combination on the second chromosome. Replicates of each of the strains were exposed to three different ethanol treatments: (i) no ethanol in the medium (control); (ii) 5% ethanol for a single generation (short-term exposure); (iii) 5% ethanol for 20 generations (long-term exposure). In all experiments, the activities of four enzymes (ADH, ODH, GPDH and AOX) were measured in larvae, pupae and adults. The results showed that (i) the larval and adult metabolic responses to environmental ethanol were different; (ii) enzyme activity changes under short-term exposure differed from those measured under long-term exposure; (iii) the activities of the allozymes common to all strains (ADH-S and GPDH-F), differed depending on the genetic background. Changes in larva-to-pupa survival were seen when the larvae of control and exposed lines of the three strains were confronted with various concentrations of ethanol. In all three strains, the exposed lines had significantly higher initial survival rate and ethanol tolerance than the control lines. Strain-specific differences were observed in the ethanol tolerance of both types of line. Received: 26 November 1996 / Accepted: 14 February 1997     

Genetic regulation of liver alcohol dehydrogenase in Peromyscus

Data from genetic crosses of Peromyscus maniculatus and P. polionotus suggest that electrophoretic variants of liver alcohol dehydrogenase are coded by alleles at a single locus. These alleles, designated Adh ^F , Adh ^S , and Adh ^N , determine, respectively, the fast, slow, and not detectable (null) ADH electrophoretic phenotypes. Heterozygotes (Adh ^F /Adh ^S ) exhibit three bands on zymograms, suggesting a dimeric subunit structure for the enzyme. However, Adh ^F /Adh ^N and Adh ^S /Adh ^N animals exhibit a single band, suggesting that the Adh ^N allele does not produce a polypeptide subunit capable of dimerizing into an active molecule. Fast and slow electrophoretic phenotypes exhibit multiple bands which can be converted into single major fast and slow bands, respectively, upon treatment with oxidized or reduced NAD. Addition of NAD also stabilizes both the fast and slow enzyme to heat inactivation at 60 C for at least 30 min.This work was supported by Predoctoral Fellowship AA-05067 from the National Institute of Alcohol Abuse and Alcoholism to K. G. B. and South Carolina Commission on Alcohol and Drug Abuse Grant 7607. Also, partial support was provided by NIH Grant CA-16184.     

Alcohol dehydrogenase polymorphism in barrel cactus populations of Drosophila mojavensis

Starch gel electrophoresis revealed that the alcohol dehydrogenase (ADH-2) locus was polymorphic in two populations (from Agua Caliente, California and the Grand Canyon, Arizona) of cactophilic Drosophila mojavensis that utilize barrel cactus (Ferocactus acanthodes) as a host plant. Electromorphs representing products of a slow (S) and a fast (F) allele were found in adult flies. The frequency of the slow allele was 0.448 in flies from Agua Caliente and 0.659 in flies from the Grand Canyon. These frequencies were intermediate to those of the low (Baja California peninsula, Mexico) and high (Sonora, Mexico and southern Arizona) frequency Adh-2^S populations of D. mojavensis that utilize different species of host cacti.     

Regulation of the activity of alcohol dehydrogenase isozymes during the development of the maize kernel

Summary The relative activities of alcohol dehydrogenase isozymes have been studied during the development of the endosperm and scutellum of heterozygous Adh ₁ ^F /Adh ₁ ^S maize kernels. The products of the Adh ₁ ^F allele are found earlier than the products of the Adh ₁ ^S allele in both the scutellum and the endosperm. A second gene (Adh _r )which controlsthe activity level of ADH is active in the scutellum only. The Adh _r ^N allele specifies increase in the relative activity of the Adh ₁ ^S products from 26 to 38 days after pollination. This increase is prevented by the Adh _r ^L allele which is dominant. These results ar discussed on the basis of the limited factor hypothesis proposed recently by Schwartz (1971) for the regulation of the Adh ₁gene in maize.     

Population genetics of the metabolically related Adh,Gpdh and Tpi polymorphisms in Drosophila melanogaster I. Geographic variation in Gpdh and Tpi allele frequencies in different continents

Among Australasian populations from above 32.5° latitude there is a significant negative relationship between Gpdh ^F frequency and distance from the equator which is not explained by gametic disequilibrium with the linked inversion In(2L)t. This is consistent with the associations reported earlier for Gpdh ^F among populations covering comparable latitudes in North America and Europe/Asia. By contrast, Tpi allele frequencies are found to be significantly associated with distance from the equator in Australasia but not North America or Europe/Asia. The Tpi pattern in the different zones is essentially the same as that reported earlier for the Acph polymorphism, which maps only 0.2 cM away from the Tpi locus.There are now ten enzyme polymorphisms in D. melanogaster which have been screened for latitudinal associations in Australasia, North America and Europe/Asia. Allele frequencies at six of these loci show significant relationships with distance from the equator which are consistent across all three zones. These latitudinal associations are more prevalent for Group II than Group I enzymes. Values of genic heterozygosity averaged over the ten polymorphic loci and eleven other monomorphic systems do not vary with latitude but differ substantially between zones. Values of Nei's genetic distance between North American and European/Asian populations calculated from all 21 systems are equivalent to subspecific differences elsewhere in the genus.     

Adaptive significance of the action of the Drosophila melanogaster alcohol dehydrogenase locus through the heat shock protein system

Four Drosophila melanogaster strains, each homozygous for one of the two major ADH aliozymes, Fast and Slow (Adh ^F1, Adh ^S , Adh ^F2 and Adh ^S2) were used to study the interaction of the Adh locus with ethanol and temperature. The separate and especially the combined effects of these two parameters allow the conclusion that the Adh locus of D. melanogaster intervenes in the adaptation process through the heat shock protein system.     

Latitudinal variation in octanol dehydrogenase and acid phosphatase allele frequencies in Drosophila melanogaster

Summary The octanol dehydrogenase (Odh) and acid phosphatase (Acph) loci of Drosophila melanogaster are each polymorphic for two electrophoretically detectable alleles. The frequencies of the Odh and Acph alleles have been analysed in populations sampled from up to a 40 ° latitudinal range in each of Australasia, North America and Europe/Asia. Odh ^S frequency is found to be significantly negatively associated with distance from the equator in all three zones. The relationship of Acph ^S frequency to distance from the equator is significant and negative in Australasia but neither significant nor consistent in sign in North America and Europe/Asia.     

The alcohol dehydrogenase alleloenzymes AdhS and AdhF from the fruitfly Drosophila melanogaster: An enzymatic rate assay to determine the active-site concentration

A rapid and reproducible enzymatic rate assay for the quantitative determination of the concentration of active sites is presented for the alleloenzymes Adh^S and Adh^F from Drosophila melanogaster. Using this procedure the turnover numbers as catalytic-center activities were found to be 12.2 sec^–1 for Adh^F and 3.4 sec^–1 for Adh^S with secondary alcohols. This showed a slower dissociation of the coenzyme from the binary enzyme-NADH complex with Adh^S and hence a stronger binding of NADH to this alleloenzyme. With ethanol, the catalytic-center activity was 1.4 sec^–1 for Adh^S and 2.8 sec^–1 for Adh^F, and hence the single amino acid mutation distinguishing the two alleloenzymes also affected hydride transfer.     

Analysis of genetic variation affecting the relative activities of fast and slow ADH dimers in Drosophila melanogaster heterozygotes

When Adh ^F /Adh ^S heterozygote homogenates are stained after electrophoresis, considerable variation is observed in the activity ratio of the FF dimer to the SS dimer. Two Adh ^S strains showed a sharp, consistent difference when crossed to a common Adh ^F strain. Optical scanning and genetic analysis confirmed that this difference originates close to the Adh locus. Since the morphs varied concordantly in their activities on numerous alcohols, and since aging and heat-treatment experiments failed to reveal a stability difference, it is proposed that the difference is regulatory in nature, affecting ADH synthesis and primarily cis-acting. A survey of wild flies revealed additional variation in the FF/SS activity ratio. Further genetic analysis showed that the basis of this variation is not restricted to the second chromosome. Furthermore, modification of the activity ratio implies some degree of allelespecificity on the part of the modifiers.This work was supported in part by money collected by Jewish Community of Iowa City, Iowa, and by NSF Grant 76-01903 to Roger Milkman.     

Perturbation of gene frequencies in a natural population of Drosophila melanogaster: evidence for selection at the Adh locus

The cellar population of Drosophila melanogaster at the Chateau Tahbilk Winery (Victoria, Australia) was perturbed for alcohol dehydrogenase (Adh) gene frequencies. Phenol oxidase (Phox) frequencies were also perturbed and monitored as a control. Subsequent gene frequency changes, together with information on population structure, indicated that selection acted on the chromosome regions of both loci. Adh gene frequencies returned to preperturbation levels in a predictable manner. A model in which the relative fitness of Adh phenotypes was determined by temperature-dependent specific activities of enzymes of Adh genotypes adequately accounts for the rate of gene frequency change at this locus. Thus temperature behaves as a selective agent in modulating Adh gene frequencies in this cellar environment.