Epistatic interactions between smell-impaired loci in Drosophila melanogaster

Odor-guided behavior is a polygenic trait determined by the concerted expression of multiple loci. Previously, P-element mutagenesis was used to identify single P[lArB] insertions, in a common isogenic background, with homozygous effects on olfactory behavior. Here, we have crossed 12 lines with these smell impaired (smi) mutations in a half-diallel design (excluding homozygous parental genotypes and reciprocal crosses) to produce all possible 66 doubly heterozygous hybrids with P[lArB] insertions at two distinct locations. The olfactory behavior of the transheterozygous progeny was measured using an assay that quantified the avoidance response to the repellent odorant benzaldehyde. There was significant variation in general combining abilities of avoidance scores among the smi mutants, indicating variation in heterozygous effects. Further, there was significant variation among specific combining abilities of each cross, indicating dependencies of heterozygous effects on the smi locus genotypes, i.e., epistasis. Significant epistatic interactions were identified for nine transheterozygote genotypes, involving 10 of the 12 smi loci. Eight of these loci form an interacting ensemble of genes that modulate expression of the behavioral phenotype. These observations illustrate the power of quantitative genetic analyses to detect subtle phenotypic effects and point to an extensive network of epistatic interactions among genes in the olfactory subgenome.     

The determination of biogenic amines in four strains of the fruit fly Drosophila melanogaster

A range of biogenic amines were measured in the heads from four strains of Drosophila melanogaster. Quantitation was carried out using gas chromatography-negative ion chemical ionization mass spectrometry (GC-NICIMS) with stable isotope dilution. The principal amines detected in the heads were dopamine, noradrenaline and 5 HT with small amounts of p- and m-tyramine; p-octopamine could not be detected in samples of 25 heads with a limit of detection of 10 pg per sample. In addition to conventional neurotransmitters or putative neurotransmitters the amines 5- and 6-hydroxydopamine were detected in the heads in substantial amounts.     

Polygenic mutation in Drosophila melanogaster: genetic interactions between selection lines and candidate quantitative trait loci

We have investigated genetic interactions between spontaneous mutations affecting abdominal and sternopleural bristle number that have accumulated in 12 long-term selection lines derived from an inbred strain, and mutations at 14 candidate bristle number quantitative trait loci. The quantitative test for complementation was to cross the selection lines to an inbred wild-type strain (the control cross) and to a derivative of the control strain into which the mutant allele at the candidate locus to be tested was substituted (the tester strain). Genetic interactions between spontaneous mutations affecting bristle number and the candidate locus mutations were common, and in several cases the interaction effects were different in males and females. Analyses of variance of the (tester- control) differences among and within groups of replicate lines selected in the same direction for the same trait showed significant group effects for several candidate loci. Genetically, the interactions could be caused by allelism of, and/ or epistasis between, spontaneous mutations in the selection lines and the candidate locus mutations. It is possible that much of the response to selection was from new mutations at candidate bristle number quantitative trait loci, and that for some of these loci, mutation rates were high.     

Papillary thyroid carcinoma oncogene (RET/PTC) alters the nuclear envelope and chromatin structure

Current evidence suggests the papillary thyroid carcinoma oncogene (RET/PTC) generates papillary thyroid carcinomas in one genetic step. We tested a resulting prediction that RET/PTC expression in thyroid epithelium should be sufficient to cause the changes in nuclear morphology diagnostic of this tumor. Primary cultures of human thyroid epithelial cells were infected with a RET/PTC retroviral construct. Morphological scoring by two independent cytopathologists shows RET/PTC expression by immunohistochemistry to be highly associated (p < 0.0001) with an irregular nuclear contour and a euchromatic appearance compared with non-expressing cells in the same cultures. The altered nuclear morphology is not due to gene transfer or transformation per se as primary thyroid cell cultures infected with a retroviral H-RAS construct differ from RET/PTC-infected cells by showing round nuclear envelopes and coarser chromatin, as determined by the independent scoring of two cytopathologists (p < 0.0001). In addition, RET/ PTC-transfected cells appear to disperse, whereas RAS-transfected cells grow as discrete colonies. The results provide additional support for the hypothesis that RET/PTC is sufficient to cause papillary thyroid carcinomas. A signaling pathway downstream of RET/ PTC leads to restructuring of the nuclear envelope and chromatin, and the signal does not depend entirely, if at all, on a RAS pathway.     

A genetic analysis of hermaphrodite, a pleiotropic sex determination gene in Drosophila melanogaster

Sex determination in Drosophila is controlled by a cascade of regulatory genes. Here we describe hermaphrodite (her), a new component of this regulatory cascade with pleiotropic zygotic and maternal functions. Zygotically, her+ function is required for female sexual differentiation: when zygotic her+ function is lacking, females are transformed to intersexes. Zygotic her+ function may also play a role in male sexual differentiation. Maternally, her+ function is needed to ensure the viability of female progeny: a partial loss of her+ function preferentially kills daughters. In addition, her has both zygotic and maternal functions required for viability in both sexes. Temperature sensitivity prevails for all known her alleles and for all of the her phenotypes described above, suggesting that her may participate in an intrinsically temperature-sensitive process. This analysis of four her alleles also indicates that the zygotic and maternal components of of her function are differentially mutable. We have localized her cytologically to 36A3-36A11.     

Population genetics and phylogenetics of DNA sequence variation at multiple loci within the Drosophila melanogaster species complex

Two regions of the genome, a 1-kbp portion of the zeste locus and a 1.1-kbp portion of the yolk protein 2 locus, were sequenced in six individuals from each of four species: Drosophila melanogaster, D. simulans, D. mauritiana, and D. sechellia. The species and strains were the same as those of a previous study of a 1.9-kbp region of the period locus. No evidence was found for recent balancing or directional selection or for the accumulation of selected differences between species. Yolk protein 2 has a high level of amino acid replacement variation and a low level of synonymous variation, while zeste has the opposite pattern. This contrast is consistent with information on gene function and patterns of codon bias. Polymorphism levels are consistent with a ranking of effective population sizes, from low to high, in the following order: D. sechellia, D. melanogaster, D.mauritiana, and D. simulans. The apparent species relationships are very similar to those suggested by the period locus study. In particular, D. simulans appears to be a large population that is still segregating variation that arose before the separation of D. mauritiana and D. sechellia. It is estimated that the separation of ancestral D. melanogaster from the other species occurred 2.5-3.4 Mya. The separations of D. sechellia and D. mauritiana from ancestral D. simulans appear to have occurred 0.58-0.86 Mya, with D. mauritiana having diverged from ancestral D. simulans 0.1 Myr more recently than D. sechellia.     

virilizer regulates Sex-lethal in the germline of Drosophila melanogaster

BACKGROUND AND PURPOSE: Advancing age is associated with declines in motor function; understanding age-related changes in the basal ganglia, therefore, is imperative for comprehension of such functional changes. The purpose of this study was to examine the age, sex, and hemispheric differences in volume of the caudate nucleus, the putamen, and the globus pallidus. METHODS: In a sample of 148 healthy right-handed adults (18-77 years old) with no evidence of age-related motor disorders, we estimated the volume of the head of the caudate nucleus, the putamen, and the globus pallidus from MR images. RESULTS: The analyses revealed bilateral age-related shrinkage of the head of the caudate nucleus and the putamen in both sexes. In men, the age-related shrinkage of the caudate was stronger on the left, whereas, in women, the opposite trend was evident. In both sexes, age-related shrinkage of the right putamen was greater than of its left counterpart. The mild bilateral age-related shrinkage of the globus pallidus was observed only in men. In both sexes, we observed significant rightward asymmetry in the putamen, significant leftward asymmetry in the caudate, and no asymmetry in the globus pallidus. CONCLUSIONS: Bilateral age-related shrinkage of the neostriatum is found in healthy adults. The shrinkage of the globus pallidus is less pronounced and may be restricted to men only.     

Formation of the male pronuclear lamina in Drosophila melanogaster

E. coli homologs of the signal recognition particle (SRP) and its receptor are essential for viability, but their role in protein export is unclear. To elucidate their function, we devised a genome-wide screen to identify genes that encode SRP substrates. Inhibition of the SRP pathway sharply blocked the membrane insertion of several polytopic inner membrane proteins (IMPs) that were predicted to be SRP substrates, but had a smaller effect on the insertion of other IMPs and no significant effect on preprotein translocation. Our results suggest that whereas most E. coli preproteins and some IMPs can utilize SRP-independent targeting pathways effectively, the structural features of a subset of IMPs have required the conservation of an SRP-based targeting machinery.     

Reduced natural selection associated with low recombination in Drosophila melanogaster

Synonymous codons are not used equally in many organisms, and the extent of codon bias varies among loci. Earlier studies have suggested that more highly expressed loci in Drosophila melanogaster are more biased, consistent with findings from several prokaryotes and unicellular eukaryotes that codon bias is partly due to natural selection for translational efficiency. We link this model of varying selection intensity to the population-genetics prediction that the effectiveness of natural selection is decreased under reduced recombination. In analyses of 385 D. melanogaster loci, we find that codon bias is reduced in regions of low recombination (i.e., near centromeres and telomeres and on the fourth chromosome). The effect does not appear to be a linear function of recombination rate; rather, it seems limited to regions with the very lowest levels of recombination. The large majority of the genome apparently experiences recombination at a sufficiently high rate for effective natural selection against suboptimal codons. These findings support models of the Hill-Robertson effect and genetic hitchhiking and are largely consistent with multiple reports of low levels of DNA sequence variation in regions of low recombination.     

Microsatellite variation in Drosophila melanogaster and Drosophila simulans: a reciprocal test of the ascertainment bias hypothesis

Interspecific comparisons of microsatellite loci have repeatedly shown that the loci are longer and more variable in the species from which they are derived (the focal species) than are homologous loci in other (nonfocal) species. There is debate as to whether this is due to directional evolution or to an ascertainment bias during the cloning and locus selection processes. This study tests these hypotheses by performing a reciprocal study. Eighteen perfect dinucleotide microsatellite loci identified from a Drosophila simulans library screen and 18 previously identified in an identical Drosophila melanogaster library screen were used to survey natural populations of each species. No difference between focal and nonfocal species was observed for mean PCR fragment length. However, heterozygosity and number of alleles were significantly higher in the focal species than in the nonfocal species. The most common allele in the Zimbabwe population of both species was sequenced for 31 of the 36 loci. The length of the longest stretch of perfect repeat units is, on average, longer in the focal species than in the non-focal species. There is a positive correlation between the length of the longest stretch of perfect repeats and heterozygosity. The difference in heterozygosity can thus be explained by a reduction in the length of the longest stretch of perfect repeats in the nonfocal species. Furthermore, flanking-sequence length difference was noted between the two species at 58% of the loci sequenced. These data do not support the predictions of the directional-evolution hypothesis; however, consistent with the ascertainment bias hypothesis, the lower variability in nonfocal species is an artifact of the microsatellite cloning and isolation process. Our results also suggest that the magnitude of ascertainment bias for repeat unit length is a function of the microsatellite size distribution in the genomes of different species.     

Molecular cloning and expression of stromalin protein from Drosophila melanogaster: homologous to mammalian stromalin family of nuclear proteins

We report the cloning of a new cDNA from Drosophila melanogaster that encodes an open reading frame of 1116 amino acid residues. It is the insect homolog of the previously reported stromalin (SA) family of nuclear proteins in mammals (Carramolino et al. [1997]. Gene 195, 151-159). Taking into account the identical domain present in all the SA family members characterized to date, we have carried out polymerase chain reaction (PCR) using degenerate oligonucleotides from the 5' and 3' ends of one of those regions of the molecule and cDNA from D. melanogaster embryos. We isolated the homologous domain of the putative Drosophila SA molecule (DSA). This cDNA fragment was used as a radiolabeled probe for screening a cDNA library from Drosophila embryos, and we have cloned a full-length cDNA for the SA homolog from an insect. The protein shows a good degree of identity with the mammalian stromalins SA-1 and SA-2, with the N and C ends being the most divergent regions of the molecule. The mRNA coding for this protein shows a molecular size of about 3.7 kb by Northern blot analysis and is essentially expressed in embryonic stage. The in situ hybridization experiments indicate that the DSA messenger is expressed mainly in neurogenic territories in the embryonic development of Drosophila. The DSA protein has been cloned and expressed in a baculovirus system, and polyclonal antibodies were generated against the recombinant molecule. Western blot analysis using these antibodies detected a main band corresponding to about 120 kDa, principally in embryos.