Electrophoretic and heat-stability polymorphism at the phosphoglucomutase (Pgm) locus in natural populations of Drosophila melanogaster

Genetic polymorphism for electrophoretic and heat-sensitive alleles is known at the phosphoglucomutase (Pgm) locus in Drosophila melanogaster. Analysis of the distribution of electrophoretic and thermosensitive (ts) alleles was carried out in natural populations from Canada and West Africa and compared with already known data on Italian populations [Trippa, G., Loverre, A., and Catamo, A. (1976). Nature 260:42]. The data show the existence of five common alleles, Pgm ^1.00,tr, Pgm ^1,00,ts, Pgm ^0.70,ts, Pgm ^1.20,ts, and Pgm ^1.50,tr, and two rare alleles, Pgm ^0.55,ts and Pgm ^1.20,tr. The most frequent allele is always Pgm ^1.00,tr; the second most common allele is always of the ts type. The cumulated frequencies of ts alleles in the populations varies between 11 and 32%. The heat stability polymorphism is present in all populations examined and shows again the uniform geographic pattern that has been found for electrophoretic variation at this locus.This research was partially supported by an operating grant (to G.R.C.) from the Canadian National Science and Engineering Research Council (NSERC).     

Genetic and biochemical studies on glutamate-pyruvate transaminase from Drosophila melanogaster

We have used electrophoretic variants of glutamate-pyruvate transaminase (GPT, E.C. 2.6.1.2) in Drosophila melanogaster to genetically map the structural gene to position 42.6 on the X chromosome. By pseudodominance tests over several deficiencies we have localized it cytogenetically to the interval 11Fl-2 to 12Al-2. The sedimentation constant (s _20,w) of the native enzyme was determined in sucrose density gradients to be 5.9 and the native molecular weight approximately 87,000. The similarity in physical properties to mammalian enzymes suggests that the enzyme may also be dimeric in D. melanogaster.     

Investigations on the PGM (phosphoglucomutase) polymorphism by isoelectric focusing inDrosophila melanogaster

Pgm allele frequencies of 383 individuals were determined in a sample ofDrosophila melanogaster from three laboratory Sardinian populations, using the techniques of standard electrophoresis, heat denaturation, and isoelectric focusing. The analysis of the progeny obtained from informative crosses showed that the isoelectric focusing patterns segregate in a Mendelian way. ThePgm ^1.00 andPgm ^0.70 electrophoretic alleles displayed different isoelectric points, whereas thePgm ^1.00,tr andPgm ^1.00,ts isoelectrophoretic alleles could not be differentiated when tested by isoelectric focusing. Moreover, thePgm ^0.70,ts allele was split into two classes, with isoelectric points ofpH 6.4 andpH 6.6.     

Enzyme polymorphism in Drosophila melanogaster populations in Iraq

Electrophoretic studies of the degree and pattern of polymorphism at two third-chromosome loci, esterase-6 (Est-6) and phosphoglucomutase (PGM), were carried out in three Drosophila melanogaster populations collected from different localities in Iraq: Mosul, Tuwaitha, and Basrah. The results show that only the Tuwaitha population was polymorphic for both loci; the other two populations were polymorphic for Est-6 and monomorphic for PGM. The allele frequency changes at both loci were followed for 20 generations in an experimental cage derived from the Tuwaitha population; it was found that there is a deviation from Hardy-Weinberg equilibrium at both loci toward the homozygote.     

Biochemical properties of esterase 6 in Drosophila melanogaster

Biochemical properties of esterase 6 in Drosophila melanogaster were investigated using partially purified preparations from three genotypes, 1/1, 1/2, and 2/2. The molecular weight of the enzyme is estimated to be about 90,000, and treatment with sodium dodecylsulfate cleaves the enzyme into four units with a molecular weight of about 22,000. The activity toward 28 naturally occurring esters was assayed and shown to vary considerably with substrate, the 1/1 preparation having in general higher activity than 1/2 and 2/2, which were very similar. Heat sensitivity, the effect of metal ions, and the effects of the presence or absence of an end product were also studied. The differences demonstrated between allozymes would allow considerable scope, under appropriate conditions, for differential selection to operate between genotypes.Supported in part by an SRC Research Studentship (N.D.D.).     

Differential characterization of two leucine aminopeptidases in Drosophila melanogaster

Two leucine aminopeptidases from Drosophila melanogaster larvae have been partially purified. The LAP A and D enzymes have similar biochemical characteristics including molecular weights of 280,000 daltons, Michaelis-Menten constants of 0.05 mM, associations with metal cofactors, and specificities toward natural and chromogenic substrates. They differ in their pH optima and spatial distributions. If the closely linked genes that code for these enzymes have resulted from a tandem gene duplication event, it is suggested that there has been subsequent evolutionary divergence. This would provide Drosophila larvae with two related, but functionally distinct enzymes.Virginia K. Walker was supported by an NRC Predoctoral Scholarship and a Killam Merit Scholarship.     

Cytogenetic localization by variation in electrophoretic allozyme phenotype: Drosophila Odh

A gene-dosage effect is characteristic of eukaryotic structural genes and is therefore useful in gene mapping. However, attributing quantitative variations in enzyme activity to a gene-dosage effect or other putative regulatory loci can be suspect when the locus in question may be inducible by variations in culture conditions. The problem of controlling for allele-specific variations in activity and regulation can be circumvented in Drosophila melanogaster by the use of synthetic duplications and deficiencies in conjunction with enzyme polymorphism. A method for constructing segmental aneupliods heterozygous for electrophoretic variants of octanol dehydrogenase (Odh) is presented which permitted variations in allozyme phenotype and enzyme activity—which show a strict dosage dependency—to be produced simultaneously. The structural gene region for Odh was identified using T(Y;A) stocks and the deficiency M(3)S31 was used to assign the locus to polytene band region 86D1–4. With this method a segmental aneuploid survey of Drosophila for purposes of gene localization can be accomplished in one generation with appropriate stocks.This work was supported by National Institutes of Health Research Grant GM18678 awarded to E. Novitski.     

Latitudinal variability of Drosophila melanogaster: Allozyme frequencies divergence between European and Afrotropical populations

Allelic frequencies at five polymorphic loci were determined in seven European and six Afrotropical populations of Drosophila melanogaster. African populations, which may be considered as ancestral for the species, showed a greater genetic diversity as measured by the number of alleles found. Within each geographic group (Europe or tropical Africa) genetic distances between local populations were very small (D=0.027). By contrast, the average distance between European and African populations (D=0.389) was more than 12 times bigger. It was previously known that various morphological or physiological differences, which probably reflect genetic adaptations to different environments, exist between these temperate and tropical populations. Data presented here suggest that the divergence in allozyme frequencies also reflects some selective mechanisms.     

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.     

Partial cytoplasmic incompatibility between two Australian populations of Drosophila melanogaster

Drosophila melanogaster (Meigen) females from stocks collected at Melbourne (latitude 37°S) show partial incompatibility when mated with males from stocks collected at Townsville (latitude 19°S) on the east coast of Australia. The reciprocal cross is compatible. Eggs have reduced hatchability in the incompatible cross. The incompatibility is maternally inherited over three generations. Compatibility can be restored by culturing Townsville flies on medium with tetracycline for one generation and by using 2-week-old Townsville males.

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Incompatibilité cytoplasmique partielle entre deux populations australiennes de Drosophila melanogaster

Résumé Les souches de D. melanogaster récoltées à Melbourne (37°S) et Townsville (19°S) sur la côte Est de l'Australie montrent une incompatibilité partielle lorsque les femelles Melbourne sont accouplées aux mâles Townsville. Une telle incompatibilité n'est décelée, ni dans les croisements intrapopulations, ni dans le croisement réciproque. Le taux d'éclosion des oeufs est réduit d'environ 30% dans le croisement incompatible, mais la viabilité des larves n'est pas modifiée. Les éléments, mâle et femelle, de ce système d'incompatibilité sont hérités maternellement pendant 3 générations de croisements en retour. La compatibilité peut être intégralement rétablie en cultivant pendant une génération la souche Townsville avec un régime contenant de la tétracycline, et partiellement rétablie en utilisant des mâles âgés de 2 semaines.

     

Correlated responses to selection for wing length in allozyme systems of Drosophila melanogaster

Summary Significant changes of genotypic structure in 20 lines selected for wing length are detected by analysis of the allelic frequencies of several enzyme loci (XDH, LAP-D, EST-6, 1-APH, ADH, -GPDH). These changes are not haphazard but a consequence of the effects of selection on the genetic structure of the population, since replicate lines always behave in a parallel way. The changes are larger in the lines selected for short wings, in which the genetic variability decreases considerably. This decrease is the result of selection for homozygosity, detected at the allozyme loci, but most probably reflects homozygosity of more or less extended chromosomal segments. Selection for wing length, especially for short wings, favoured recombinants of the initial founder chromosomes. Only in the 1-APH and the EST-6 loci, separated by 11.7 centimorgans on the genetic map, do the alleles linked in the founder lines change in parallel in the control and long wing lines. The correlated response in the allozyme allele frequencies cannot be accounted for by a direct influence of the allozymes on the variability in wing length. The changes in the EST-6, 1-APH and perhaps in the LAP-D, can be explained by a direct effect of natural selection on the allozyme loci, probably in interaction with the effect of selection for wing length on linked loci. This last effect seems to be the main factor contributing to the change detected in the XDH locus.     

DNA-dependent RNA polymerases from Drosophila melanogaster adults: Isolation and partial characterization

In preparation for the isolation and biochemical characterization of putative RNA polymerase mutants, DNA-dependent RNA polymerases of Drosophila melanogaster adults were isolated and partially characterized. Approximately 70% of the female adult RNA polymerase is located in ovaries. Multiple forms of ovarian RNA polymerases I and II are separable by DEAE-Sephadex chromatography. The two forms of RNA polymerase II differ in ammonium sulfate optima. RNA polymerase IIA is more active with double-stranded DNA as template, whereas RNA polymerase IIB transcribes single-stranded DNA most efficiently. Rechromatography of RNA polymerase IIA on DEAE-Sephadex results in the loss of ability of this form to transcribed double-stranded DNA most efficiently. Ovariectomized carcasses have two forms of RNA polymerase I and one form of RNA polymerase II and each transcribes single-stranded DNA most efficiently. As judged by gel filtration chromatography, female adult extracts have forms of RNA polymerase II that differ in molecular weight and template preference.Supported by Grants GM23456 from the NIH and 11259 from the City University Research Foundation.     

Comparative studies of allozyme loci in Drosophila simulans and Drosophila melanogaster. I. Three dipeptidase loci

Genetic variation at three dipeptidase loci (Dip-A, Dip-B, and Dip-C) in Drosophila simulans was analyzed by starch gel electrophoresis. Dip-A was found to be polymorphic in four populations, while Dip-B and Dip-C were found to be polymorphic in one. The numbers of different alleles found at each respective locus were: Dip-A, two; Dip-B, two; and Dip-C, three. Dip-A was genetically mapped at 57.9 on the second chromosome, and Dip-B and Dip-C at 80.9 and 87.9 on the third chromosome, respectively. Neither Dip-B nor Dip-C has been mapped in D. melanogaster because both loci are apparently monomorphic. Their map positions in D. simulans with respect to flanking markers whose homologous genes have been cytogenetically localized in D. melanogaster suggested that they might be mapped cytogenetically by using available deficiencies in D. melanogaster. Accordingly, by the construction of interspecific hybrids which carried deficiencies of melanogaster and an allele of simulans with a mobility different from that of the fixed melanogaster allele, Dip-B and Dip-C were localized between 87F12-14 and 88C1-3 and between 87B5-6 and 87B8-10, respectively, in the salivary gland chromosomes of D. melanogaster. The similarity between these two species is discussed on the basis of these findings.     

Clonal analysis in hybrids between Drosophila melanogaster and Drosophila simulans

We have analysed the viability of cellular clones induced by mitotic recombination in Drosophila melanogaster/D. simulans hybrid females during larval growth. These clones contain a portion of either melanogaster or simulans genomes in homozygosity. Analysis has been carried out for the X and the second chromosomes, as well as for the 3L chromosome arm. Clones were not found in certain structures, and in others they appeared in a very low frequency. Only in abdominal tergites was a significant number of clones observed, although their frequency was lower than in melanogaster abdomens. The bigger the portion of the genome that is homozygous, the less viable is the recombinant melano-gaster/simulans hybrid clone. The few clones that appeared may represent cases in which mitotic recombination took place in distal chromosome intervals, so that the clones contained a small portion of either melanogaster or simulans chromosomes in homozygosity. Moreover, Lhr, a gene of D. simulans that suppresses the lethality of male and female melanogaster/simulans hybrids, does not suppress the lethality of the recombinant melanogaster/simulans clones. Thus, it appears that there is not just a single gene, but at least one per tested chromosome arm (and maybe more) that cause hybrid lethality. Therefore, the two species, D. melanogaster and D. simulans, have diverged to such a degree that the absence of part of the genome of one species cannot be substituted by the corresponding part of the genome of the other, probably due to the formation of co-adapted gene complexes in both species following their divergent evolution after speciation. The disruption of those coadapted gene complexes would cause the lethality of the recombinant hybrid clones.     

Genetic Variants Affecting Phenoloxidase Activity in Drosophila melanogaster

In Drosophila melanogaster, two new variants affecting the activity of phenoloxidase were found in natural populations at Gomel in Belorussia and at Krasnodar in Russia. Prophenoloxidases, A ₁ and A ₃ , in these variants had the same mobilities on native electrophoresis as the wild type. However, enzymatic activities in their activated states were much lower than in the wild type, whereas the existence of prophenoloxidase proteins was demonstrated. Egg-to-adult and relative viabilities in the variants did not decrease at temperatures between 18 and 29°C. Genetic analyses indicated that the genes showing the phenotype of variants are new alleles of Mox and Dox-3 on the second chromosome.     

Biological and Behavioral Effects of Heavy Metals in Drosophila melanogaster Adults and Larvae

Heavy metals are essential components of biological systems but are extremely toxic at high doses. As a result, we hypothesized that perception of heavy metals through gustation may exist in Drosophila melanogaster. In this study, we investigated the behavioral effects of iron, copper, zinc, and cadmium on D. melanogaster gustation, oviposition, and pupation-site selection. In addition, we examined the biological effects of heavy metals on the fruit fly survival and reproductive success. Our results illustrate that D. melanogaster responds behaviorally to the presence of high concentrations of heavy metals in food. All metals acted as repellents to the fruit flies at high doses, with the egg-laying and feeding of the female flies significantly decreasing. Furthermore, supplementation of heavy metals in the culture medium reduced survival to the adult stage and shortened the life span of adult flies. From these observations, we speculate that D. melanogaster avoidance behavior towards high concentrations of heavy metals may have a positive effect on their survival and reproductive success in nature, particularly in the presence of metal-contaminated food sources.     

A simple approach for discovering common nonelectrophoretic enzyme variability: A heat denaturation study in Drosophila melanogaster

A simple procedure is described to detect genetic heterogeneity within electrophoretic classes at a locus in Drosophila, based on electrophoresis and heat denaturation studies. Temperature-resistant (tr) and temperature-sensitive (ts) isoelectrophoretic alleles at the phosphoglucomutase locus (Pgm) are present at polymorphic frequencies in natural and in laboratory populations of Drosophila melanogaster.This work has been supported by a grant from the Consiglio Nazionale delle Ricerche (C.N.R.).     

Glycomic studies of Drosophila melanogaster embryos

With the complete genome sequence of Drosophila melanogaster defined a systematic approach towards understanding the function of glycosylation has become possible. Structural assignment of the entire Drosophila glycome during specific developmental stages could provide information that would shed further light on the specific roles of different glycans during development and pinpoint the activity of certain glycosyltransferases and other glycan biosynthetic genes that otherwise might be missed through genetic analyses. In this paper the major glycoprotein N- and O-glycans of Drosophila embryos are described as part of our initial undertaking to characterize the glycome of Drosophila melanogaster. The N-glycans are dominated by high mannose and paucimannose structures. Minor amounts of mono-, bi- and tri-antennary complex glycans were observed with GlcNAc and Galβ1–4GlcNAc non-reducing end termini. O-glycans were restricted to the mucin-type core 1 Galβ1-3GalNAc sequence.     

Partial purification and properties of guanosine triphosphate cyclohydrolase from Drosophila melanogaster

The first enzyme (named GTP cyclohydrolase) in the pathway for the biosynthesis of pteridines has been partially purified from extracts of late pupae and young adults of Drosophila melanogaster. This enzyme catalyzes the hydrolytic removal from GTP of carbon 8 as formate and the synthesis of 2-amino-4-hydroxy-6-(d-erythro-1,2,3-trihydroxypropyl)-7,8-dihydropteridine triphosphate (dihydroneopterin triphosphate). Some of the properties of the enzyme are as follows: it functions optimally at pH 7.8 and at 42 C; activity is unaffected by KCl and NaCl, but divalent cations (Mg²⁺, Mn²⁺, Zn²⁺, and Ca²⁺) are inhibitory; the K _m for GTP is 22 m; and the molecular weight is estimated at 345,000 from gel filtration experiments. Of a number of nucleotides tested, only GDP and dGTP were used to any extent as substrate in place of GTP, and these respective compounds were used only 1.8% and 1.5% as well as GTP.This work was supported by research grants from the National Institutes of Health (AM03442) and the National Science Foundation (GB33929).