Courtship Anomalies Caused by Doublesex Mutations in Drosophila Melanogaster

The role played by the sex-determining gene doublesex (dsx) and its influence on Drosophila courtship were examined. Against a background of subnormal male-like behavior that is reported to be an attribute of haplo-X flies homozygous for the original dsx mutation, and given that a sex-specific muscle is unaffected by genetic variation at this locus, analyses of several reproductive behaviors and control for genetic background effects indicated that XY dsx mutants are impaired in their willingness to court females. When they did court, certain behavioral actions were normal, including components of courtship song. However, these mutants never produced courtship humming sounds. Mature XY dsx flies elicited anomalously high levels of courtship; that this occurs merely because of a delay in imaginal development was experimentally discounted. The current analysis reconciled two ostensibly conflicting reports involving the courtship-stimulating qualities of this mutant type. Such experiments also uncovered a new behavioral anomaly: dsx mutations caused chromosomal males to court other males at abnormally high levels. These results are discussed from the perspective of doublesex's influence on internal tissues of adult Drosophila involved in the triggering and neural control of male- and female-like elements of courtship, reproductive pheromone production, or a combination of such factors.     

Sequence Analysis of N-Ethyl-N-Nitrosourea-Induced Vermilion Mutations in Drosophila Melanogaster

The mutational specificity of N-ethyl-N-nitrosourea (ENU) was determined in Drosophila melanogaster using the vermilion locus as a target gene. 25 mutants (16 F1 and 9 F2 mutants) were cloned and sequenced. Only base-pair changes were observed; three of the mutants represented double base substitutions. Transition mutations were the most prominent sequence change: 61% were GC----AT and 18% AT----GC substitutions. Both sequence changes can be explained by the miscoding properties of the modified guanine and thymine bases. A strong bias of neighboring bases on the occurrence of the GC----AT transitions or a strand preference of both types of transition mutations was not observed. The spectrum of ENU mutations in D. melanogaster includes a significant fraction (21%) of transversion mutations. Our data indicate that like in other prokaryotic and eukaryotic systems also in D. melanogaster the O6-ethylguanine adduct is the most prominent premutational lesion after ENU treatment. The strong contribution of the O6-ethylguanine adduct to the mutagenicity of ENU possibly explains the absence of distinct difference between the type of mutations observed in the F1 and F2 mutants. Although the latter arise later during development, the spectrum of mosaic mutations is also dominated by GC----AT transition mutations.     

Developmental Genetic Analysis of Contrabithorax Mutations in Drosophila Melanogaster

A developmental analysis of the Contrabithorax (Cbx) alleles offers the opportunity to examine the role of the Ultrabithorax (Ubx) gene in controlling haltere, as alternative to wing, morphogenesis in Drosophila. Several Cbx alleles are known with different spatial specificity in their wing toward haltere homeotic transformation. The molecular data on these mutations, however, does not readily explain differences among mutant phenotypes. In this work, we have analyzed the "apogenetic" mosaic spots of transformation in their adult phenotype, in mitotic recombination clones and in the spatial distribution of Ubx proteins in imaginal discs. The results suggest that the phenotypes emerge from early clonality in some Cbx alleles, and from cell-cell interactions leading to recruitment of cells to Ubx gene expression in others. We have found, in addition, mutual interactions between haltere and wing territories in pattern and dorsoventral symmetries, suggesting short distance influences, "accommodation," during cell proliferation of the anlage. These findings are considered in an attempt to explain allele specificity in molecular and developmental terms.     

Molecular Analysis of Ems-Induced Frizzled Mutations in Drosophila Melanogaster

The frizzled (fz) gene of Drosophila is essential for the development of normal tissue polarity in the adult cuticle of Drosophila. In fz mutants the parallel array of hairs and bristles that decorate the cuticle is disrupted. Previous studies have shown that fz encodes a membrane protein with seven putative transmembrane domains, and that it has a complex role in the development of tissue polarity, as there exist both cell-autonomous and cell nonautonomous alleles. We have now examined a larger number of alleles and found that 15 of 19 alleles display cell nonautonomy. We have examined these and other alleles by Western blot analysis and found that most fz mutations result in altered amounts of Fz protein, and many also result in a Fz protein that migrates aberrantly in SDS-PAGE. We have sequenced a subset of these alleles. Cell nonautonomous fz alleles were found to be associated with mutations that altered amino acids in all regions of the Fz protein. Notably, the four cell-autonomous mutations were all in a proline residue located in the presumptive first cytoplasmic loop of the protein. We have also cloned and sequenced the fz gene from D. virilis. Conceptual translation of the D. virilis open reading frame indicates that the Fz protein is unusually well conserved. Indeed, in the putative cytoplasmic domains the Fz proteins of the two species are identical.     

A Functional Analysis of Notch Mutations in Drosophila

The Notch gene encodes a receptor protein that is involved in many processes during development. Its best understood role is during neurogenesis in a process called ``lateral inhibition.' However, it has been proposed that Notch also has a role in defining the proneural clusters in the first place. This raises the possibility that the Notch protein is acting as a multifunctional receptor. To test this hypothesis, we have carried out a genetic analysis of molecularly characterized Notch alleles to identify alleles that affect only one of the two proposed functions. Here we present evidence that Notch alleles can be identified that appear to affect the function of Notch during either lateral inhibition or the definition of proneural clusters. In addition our results indicate that there may be discrete regions of the Notch protein required for each function.     

Autosomal Mutations Affecting Adhesion between Wing Surfaces in Drosophila Melanogaster

Integrins are evolutionarily conserved transmembrane α,β heterodimeric receptors involved in cell-to-matrix and cell-to-cell adhesions. In Drosophila the position-specific (PS) integrins mediate the formation and maintenance of junctions between muscle and epidermis and between the two epidermal wing surfaces. Besides integrins, other proteins are implicated in integrin-dependent adhesion. In Drosophila, somatic clones of mutations in PS integrin genes disrupt adhesion between wing surfaces to produce wing blisters. To identify other genes whose products function in adhesion between wing surfaces, we conducted a screen for autosomal mutations that produce blisters in somatic wing clones. We isolated 76 independent mutations in 25 complementation groups, 15 of which contain more than one allele. Chromosomal sites were determined by deficiency mapping, and genetic interactions with mutations in the β(PS) integrin gene myospheroid were investigated. Mutations in four known genes (blistered, Delta, dumpy and mastermind) were isolated. Mutations were isolated in three new genes (piopio, rhea and steamer duck) that affect myo-epidermal junctions or muscle function in embryos. Mutations in three other genes (kakapo, kiwi and moa) may also affect cell adhesion or muscle function at hatching. These new mutants provide valuable material for the study of integrin-dependent cell-to-cell adhesion.     

Molecular Analysis of Mutations Induced in the Vermilion Gene of Drosophila Melanogaster by Methyl Methanesulfonate

The nature of DNA sequence changes induced by methyl methanesulfonate (MMS) at the vermilion locus of Drosophila melanogaster was determined after exposure of postmeiotic male germ cell stages. MMS is a carcinogen with strong preference for base nitrogen alkylation (s = 0.86). The spectrum of 40 intralocus mutations was dominated by AT----GC transitions (23%), AT----TA transversions (54%) and deletions (14%). The small deletions were preferentially found among mutants isolated in the F1 (8/18), whereas the AT----GC transitions exclusively occurred in the F2 (6/22). The MMS-induced transversions and deletions are presumably caused by N-methyl DNA adducts, which may release apurinic intermediates, known to be a time-related process. Furthermore, MMS produces multilocus deletions, i.e., at least 30% of the F1 mutants analyzed were of this type. A comparison of the mutational spectra of MMS with that produced by ethylnitrosourea (ENU), also in the vermilion locus of Drosophila, reveals major differences: predominantly transition mutations (61% GC----AT and 18% AT----GC) were found in both the F1 and F2 spectrum induced by ENU. It is concluded that the mutational spectrum of MMS is dominated by nitrogen DNA adducts, whereas with ENU DNA sequence changes mainly arose from modified oxygen in DNA.     

Competition between Delta and the Abruptex domain of Notch

Background  

Extracellular domains of the Notch family of signalling receptors contain many EGF repeat domains, as do their major ligands. Some EGF repeats are modified by O-fucosylation, and most have no identified role in ligand binding.     

Interactions of Vestigial and Scabrous with the Notch Locus of Drosophila Melanogaster

Interactions are described between the Notch locus of Drosophila melanogaster, and two other loci, scabrous and vestigial, which respectively affect the eyes and wings. The Notch locus is responsible for mediating decisions of cell fate throughout development in many different tissues. Mutations and duplications of vestigial and scabrous alter the severity of phenotypes associated with Notch mutations and duplications in a manner that is essentially tissue- and allele-specific. These interactions indicate that the products of vestigial and scabrous act in conjunction with Notch to stimulate the differentiation of specific cell types.     

P Transposon-Induced Dominant Enhancer Mutations of Position-Effect Variegation in Drosophila Melanogaster

P transposon induced modifier mutations of position-effect variegation (PEV) were isolated with the help of hybrid dysgenic crosses (π2 strain) and after transposition of the mutator elements pUChsneory(+) and P[lArB]. Enhancer mutations were found with a ten times higher frequency than suppressors. The 19 pUChsneory(+)- and 15 P[lArB]-induced enhancer mutations can be used for cloning of genomic sequences at the insertion sites of the mutator elements via plasmid rescue. Together with a large sample of X-ray-induced (48) and spontaneous (93) enhancer mutations a basic genetic analysis of this group of modifier genes was performed. On the basis of complementation and mapping data we estimate the number of enhancer genes at about 30 in the third chromosome and between 50 and 60 for the whole autosome complement. Therefore, enhancer of PEV loci are found in the Drosophila genome as frequently as suppressor genes. Many of the enhancer mutations display paternal effects consistent with the hypothesis that some of these mutations can induce genomic imprinting. First studies on the developmentally regulated gene expression of PEV enhancer genes were performed by β-galactosidase staining in P[lArB] induced mutations.     

Age-specific Effects of Novel Mutations in Drosophila Melanogaster I. Mortality

Theories for the evolution of aging rest on the assumption that at least some deleterious mutations have effects that are limited to certain ages. Many mutation accumulation studies have tried to measure the number and magnitude of deleterious mutations, but few studies have tried to determine the extent to which the effects of mutations are limited to particular ages. Here we estimate the age-specific effect of deleterious mutations on mortality rate in an outbred population of the fruit fly, Drosophila melanogaster. We used the ‘middle class neighborhood’ approach to accumulation mutations in populations of flies that had recently been obtained from the wild. There are mutations that increase mortality rates, but whose effects are limited to specific ages. The age-specificity of mutational effects differs between the sexes, between virgin and mated flies, and over time. After 10 and 20 generations of mutation accumulation, there were clear age-specific effects of mutations. After 30 generations, however, the degree of age-specificity decreased. In addition, mutation accumulation led to a steady increase in larval mortality and a small but significant increase in the sex ratio of eclosing flies. We discuss the implications of these results for models of aging, and suggest approaches that future studies should take to obtain accurate information on the age-specificity of novel mutations. This revised version was published online in July 2006 with corrections to the Cover Date.     

Cytoplasmic Dynein Function Is Essential in Drosophila Melanogaster

The microtubule motor cytoplasmic dynein has been implicated in a variety of intracellular transport processes. We previously identified and characterized the Drosophila gene Dhc64C, which encodes a cytoplasmic dynein heavy chain. To investigate the function of the cytoplasmic dynein motor, we initiated a mutational analysis of the Dhc64C dynein gene. A small deletion that removes the chromosomal region containing the heavy chain gene was used to isolate EMS-induced lethal mutations that define at least eight essential genes in the region. Germline transformation with a Dhc64C transgene rescued 16 mutant alleles in the single complementation group that identifies the dynein heavy chain gene. All 16 alleles were hemizygous lethal, which demonstrates that the cytoplasmic dynein heavy chain gene Dhc64C is essential for Drosophila development. Furthermore, our failure to recover somatic clones of cells homozygous for a Dhc64C mutation indicates that cytoplasmic dynein function is required for cell viability in several Drosophila tissues. The intragenic complementation of dynein alleles reveals multiple mutant phenotypes including male and/or female sterility, bristle defects, and defects in eye development.