A Cytogenetic Analysis of Chromosomal Region 31 of Drosophila Melanogaster

Cytogenetic region 31 of the second chromosome of Drosophila melanogaster was screened for recessive lethal mutations. One hundred and thirty nine new recessive lethal alleles were isolated that fail to complement Df(2L)J2 (31A-32A). These new alleles, combined with preexisting mutations in the region, define 52 complementation groups, 35 of which have not previously been described. Among the new mutations were alleles of the cdc2 and mfs(2)31 genes. Six new deficiencies were also isolated and characterized identifying 16 deficiency subintervals within region 31. The new deficiencies were used to further localize three loci believed to encode non-histone chromosomal proteins. Suvar(2)1/Su(var)214, a dominant suppressor of position-effect variegation (PEV), maps to 31A-B, while the recessive suppressors of PEV mfs(2)31 and wdl were localized to regions 31E and 31F-32A, respectively. In addition, the cytological position of several mutations that interact with heterochromatin were more precisely defined.     

Genetic Analysis of Chromosome Region 63 of Drosophila Melanogaster

The salivary chromosome region including cytological division 63 of Drosophila melanogaster was genetically analyzed in order to (1) characterize this previously unstudied region and (2) attempt to isolate mutations in the hsp82 gene. Seven deletions which span this region were isolated, including four which remove the hsp82 gene. A Minute mutation was mapped to this region and this Minute was used to isolate duplications in the 63 region. These duplications map the Minute to 63B8-C1. F2 screens were initiated using deletions which remove the hsp82 gene. Over 15,000 chromosomes were screened, yielding 40 lethal mutations which comprise 14 complementation groups. Several of these mutations map outside the 63 region and appear to give second site interaction with the Minute locus. Four loci, including the Minute gene, are candidates for hsp82 mutations by cytogenetic mapping. These loci were tested for complementation with a P element carrying the hsp82 gene. However, none of the mutations was rescued.     

A Genetic and Molecular Analysis of the 46c Chromosomal Region Surrounding the Fmrfamide Neuropeptide Gene in Drosophila Melanogaster

We have analyzed the FMRFamide neuropeptide gene region of Drosophila melanogaster. This gene maps to the 46C region of chromosome 2R; this interval previously was not well characterized. For this genetic and molecular analysis, we have used X-ray mutagenesis, EMS mutagenesis, and the recently reported local P element transposition method. We identified four overlapping deletions, two of which have proximal breakpoints that define a 50-60-kb region surrounding the FMRFamide gene in 46C. To this small region, we mapped three lethal complementation groups; 10 additional lethal complementation groups were mapped to more distal regions of 46CD. One of these groups corresponds to even-skipped, the other 12 are previously unidentified. Using various lines of evidence we excluded the possibility that FMRFamide corresponds to any of the three lethal complementation groups mapping to its immediate 50-60-kb vicinity. The positions of two of the three lethal complementation groups were identified with P elements using a local transposition scheme. The third lethal complementation group was excluded as being FMRFamide mutants by sequence analysis and by immunocytochemistry with proFMRFamide precursor-specific antibodies. This analysis has (1) provided a genetic map of the 46CD chromosomal region and a detailed molecular map of a portion of the 46C region and (2) provided additional evidence of the utility of local transposition for targeting nearby genes.     

Gross Genetic Dissection and Interaction of the Chromosomal Region 95e;96f of Drosophila Melanogaster

Making use of deficiencies, inversions and translocations, we have genetically dissected the region 95E to 96F of Drosophila melanogaster. We localized cytologically the loci abnormal spindle (asp: 3-85.2: 96A20-25;96B1-10) and M(3)96C2 (96C1;96C5). We have also found several new phenotypes associated with lesions in the 95E to 97B region: (1) Minute(3)96A (M(3)96A) is a haplo-insufficient phenotype of thin and short bristles presented by individuals deficient for the region 95E6-8;96A1-5. (2) abdominal-one reduced (aor) shows two different phenotypes associated with the distal breakpoint of In(3R)Ubx7L (89E;96A1-7). One is the increase of the Ubx phenotype, but its effect requires the presence of lesions in Ubx. The other phenotype is a drastic reduction or disappearance of the first abdominal segment. Both phenotypes might be due to lesions in the same gene. (3) metaphase arrest (mar) is associated with the breakpoint of the T(Y;3)B197 (96B1-10) and produces a phenotype typical of mitotic mutants with arrest of the cell cycle during prometaphase or metaphase. There is another region localized in 97B which interacts with asp: in a background homozygous for asp, three doses of this region enhance the asp phenotype.     

Genetic Analysis of Stellate Elements of Drosophila Melanogaster

Repeated elements are remarkably important for male meiosis and spermiogenesis in Drosophila melanogaster. Pairing of the X and Y chromosomes is mediated by the ribosomal RNA genes of the Y chromosome and X chromosome heterochromatin, spermiogenesis depends on the fertility factors of the Y chromosome. Intriguingly, a peculiar genetic system of interaction between the Y-linked crystal locus and the X-linked Stellate elements seem to be also involved in male meiosis and spermiogenesis. Deletion of the crystal element of the Y, via an interaction with the Stellate elements of the X, causes meiotic abnormalities, gamete-genotype dependent failure of sperm development (meiotic drive), and deposition of protein crystals in spermatocytes. The current hypothesis is that the meiotic abnormalities observed in cry(-) males is due to an induced overexpression of the normally repressed Ste elements. An implication of this hypothesis is that the strength of the abnormalities would depend on the amount of the Ste copies. To test this point we have genetically and cytologically examined the relationship of Ste copy number and organization to meiotic behavior in cry(-) males. We found that heterochromatic as well as euchromatic Ste repeats are functional and that the abnormality in chromosome condensation and the frequency of nondisjunction are related to Ste copy number. Moreover, we found that meiosis is disrupted after synapsis and that cry-induced meiotic drive is probably not mediated by Ste.     

In Situ Hybridization Analysis of Chromosomal Homologies in Drosophila Melanogaster and Drosophila Virilis

Twenty-four biotin-labeled recombinant-DNA probes which contained putative unique-sequence Drosophila melanogaster DNA were hybridized to larval salivary-gland chromosomes of D. melanogaster and Drosophila virilis. All probes hybridized to D. melanogaster chromosomes at the expected sites. However, one probe hybridized to at least 16 additional sites, and one hybridized to one additional site. Thirteen probes hybridized strongly to D. virilis chromosomes, four hybridized weakly and infrequently, and seven did not hybridize. Probes representing two multigene families (beta-tubulin and yolk-protein) hybridized as would be expected if all sites had been conserved in the two species on the same chromosomal elements. The multiple hybridization sites of a third probe which may represent a multigene family were also conserved. The results were consistent with H.J. Muller's proposal that chromosomal elements have been conserved during evolution of this genus.     

A Genetic Analysis of Male-Predominant Pheromones in Drosophila Melanogaster

Chemical signals from males play an important role in stimulating Drosophila melanogaster females to mate, and male-predominant pheromones may influence a female's choice of mates. Male-predominant pheromones also inhibit courtship, thereby functioning as antiaphrodisiacs. Interstrain variation in the ratio of two male-predominant pheromones (7-tricosene and 7-pentacosene) has been reported, but the genetic basis for this potentially important variation has not been examined. In a series of crosses between strains that differ radically in the amounts of 7-tricosene and 7-pentacosene, we have identified both X-linked and autosomal contributions to interstrain variation in the amounts of these compounds. The X-linked loci act as enhancers for production of the compound predominant in the strain from which the X chromosome originated. Autosomal factors for each of the two compounds appear to segregate as high vs. low, with incomplete dominance of high 7-tricosene over low, and low 7-pentacosene over high. A significant negative correlation between the quantities of 7-pentacosene and 7-tricosene in the F2 and backcross progeny, but not in the F1s or parentals, indicates linkage between autosomal loci regulating the expression of each compound. However, the phenotypic distributions of the backcross progeny indicate that additional unlinked loci are also directly involved in the production of these two hydrocarbons.     

Genetic Analysis of the Claret Locus of Drosophila Melanogaster

The claret (ca) locus of Drosophila melanogaster comprises two separately mutable domains, one responsible for eye color and one responsible for proper disjunction of chromosomes in meiosis and early cleavage divisions. Previously isolated alleles are of three types: (1) alleles of the claret (ca) type that affect eye color only, (2) alleles of the claret-nondisjunctional (cand) type that affect eye color and chromosome behavior, and (3) a meiotic mutation, non-claret disjunctional (ncd), that affects chromosome behavior only. In order to investigate the genetic structure of the claret locus, we have isolated 19 radiation-induced alleles of claret on the basis of the eye color phenotype. Two of these 19 new alleles are of the cand type, while 17 are of the ca type, demonstrating that the two domains do not often act as a single target for mutagenesis. This suggests that the two separately mutable functions are likely to be encoded by separate or overlapping genes rather than by a single gene. One of the new alleles of the cand type is a chromosome rearrangement with a breakpoint at the position of the claret locus. If this breakpoint is the cause of the mutant phenotype and there are no other mutations associated with the rearrangement, the two functions must be encoded by overlapping genes.     

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.     

Genetic and Molecular Mapping of Chromosome Region 85a in Drosophila Melanogaster

Chromosome region 85A contains at least 12 genetic complementation groups, including the genes dhod, pink and hunchback. In order to better understand the organization of this chromosomal segment and to permit molecular studies of these genes, we have carried out a genetic analysis coupled with a chromosome walk to isolate the DNA containing these genes. Complementation tests with chromosomal deficiencies permitted unambiguous ordering of most of the complementation groups identified within the 85A region. Recombinant bacteriophage clones were isolated that collectively span over 120 kb of 85A DNA and these were used to produce a molecular map of the region. The breakpoint sites of a number of 85A chromosome rearrangements were localized on the molecular map, thereby delimiting regions of the DNA that contain the various genetic complementation groups.     

Cytogenetic Analysis of Chromosome Region 73ad of Drosophila Melanogaster

The 73AD salivary chromosome region of Drosophila melanogaster was subjected to mutational analysis in order to (1) generate a collection of chromosome breakpoints that would allow a correlation between the genetic, cytological and molecular maps of the region and (2) define the number and gross organization of complementation groups within this interval. Eighteen complementation groups were defined and mapped to the 73A2-73B7 region, which is comprised of 17 polytene bands. These complementation groups include the previously known scarlet (st), transformer (tra) and Dominant temperature-sensitive lethal-5 (DTS-5) genes, as well as 13 new recessive lethal complementation groups and one male and female sterile locus. One of the newly identified lethal complementation groups corresponds to the molecularly identified abl locus, and another gene is defined by mutant alleles that exhibit an interaction with the abl mutants. We also recovered several mutations in the 73C1-D1.2 interval, representing two lethal complementation groups, one new visible mutant, plucked (plk), and a previously known visible, dark body (db). There is no evidence of a complex of sex determination genes in the region near tra.     

Somatic Reversion of Chromosomal Position Effects in Drosophila Melanogaster

A transgene was inserted at several different chromosomal sites in Drosophila melanogaster, where its expression was subject to genomic position effects. Quantitative position effects and variegated and constant patterned position effects were observed. We investigated the status of the affected gene in the somatic cells where it normally functions. The FLP site-specific recombinase was used to remove the gene from the chromosome and its expression was then evaluated. We show that the FLP recombinase functions in cells that have finished their developmental program of mitoses. When FLP acts on directly repeated copies of its target site (FRT), the DNA flanked by those FRTs is excised from the chromosome as a closed circle. The extrachromosomal circle is maintained in nondividing cells, and a gene located on such a circle can be expressed. We then demonstrate that a gene subject to either variegated or constant position effect can be relieved of that effect by excision of the gene from the chromosome in cells where it would otherwise be inactive. We also observed a strong inhibition of FLP-mediated recombination for target sites located near centric heterochromatin.     

Genetic Analysis of the Shaker Gene Complex of Drosophila Melanogaster

The Shaker complex (ShC) spans over 350 kb in the 16F region of the X chromosome. It can be dissected by means of aneuploids into three main sections: the maternal effect (ME), the viable (V) and the haplolethal (HL) regions. The mutational analysis of ShC shows a high density of antimorphic mutations among 12 lethal complementation groups in addition to 14 viable alleles. The complex is the structural locus of a family of potassium channels as well as a number of functions relevant to the biology of the nervous system. The constituents of ShC seem to be linked by functional relationships in view of the similarity of the phenotypes, antimorphic nature of their mutations and the behavior in transheterozygotes. We discuss the relationship between the genetic organization of ShC and the functional coupling of potassium currents with the other functions encoded in the complex.     

Genetic Analysis of the Adenosine3 (Gart) Region of the Second Chromosome of Drosophila Melanogaster

The Gart gene of Drosophila melanogaster is known, from molecular biological evidence, to encode a polypeptide that serves three enzymatic functions in purine biosynthesis. It is located in polytene chromosome region 27D. One mutation in the gene (ade3(1)) has been described previously. We report here forty new ethyl methanesulfonate-induced mutations selected aga!nst a synthetic deficiency of the region from 27C2-9 to ++28B3-4. The mutations were characterized cytogenetically and by complementation analysis. The analysis apparently identifies 12 simple complementation groups. In addition, two segments of the chromosome exhibit complex complementation behavior. The first, the 28A region, gave three recessive lethals and also contains three known visible mutants, spade (spd), Sternopleural (Sp) and wingless (wg); a complex pattern of genetic interaction in the region incorporates both the new and the previously known mutants. The second region is at 27D, where seven extreme semilethal mutations give a complex complementation pattern that also incorporates ade3(1). Since ade3(1) is defective in one of the enzymatic functions encoded in the Gart gene, we assume the other seven also affect the gene. The complexity of the complementation pattern presumably reflects the functional complexity of the gene product. The phenotypic effects of the mutants at 27D are very similar to those described for ade2 mutations, which also interrupt purine biosynthesis.     

Genetic Analysis of the Additional Sex Combs Locus of Drosophila Melanogaster

Additional sex combs (Asx) is a member of the Polycomb group of genes, which are thought to be required for maintenance of chromatin structure. To better understand the function of Asx, we have isolated nine new alleles, each of which acts like a gain of function mutation. Asx is required for normal determination of segment identity. AsxP1 shows an unusual phenotype in that anterior and posterior homeotic transformations are seen in the same individuals, suggesting that AsxP1 might upset chromatin structure in a way that makes both activation and repression of homeotic genes more difficult. Analysis of embryonic and adult phenotypes of Asx alleles suggests that Asx is required zygotically for determination of segment number and polarity. The expression pattern of even-skipped is altered in Asx mutant embryos, suggesting that Asx is required for normal expression of this gene. We have transposon-tagged the Asx gene, and can thus begin molecular analysis of its function.