Cytogenetic Analysis of Chromosome 3 in DROSOPHILA MELANOGASTER: Mapping of the Proximal Portion of the Right Arm

In order to define more precisely the most proximal portion of chromosome 3R in Drosophila melanogaster, several new chromosome aberrations involving this region have been recovered and analyzed. These new arrangements were recovered as induced reversions of two dominant mutations, Antp^Ns and dsx^D, located in the region of interest. The results of the analysis have allowed the localization of several existing mutations, have further elucidated the complex homoeotic locus which resides in this region, and have confirmed the efficacy of this type of screen in the analysis of specific chromosome regions.     

On the Components of Segregation Distortion in DROSOPHILA MELANOGASTER

The segregation distorter (SD) complex is a naturally occurring meiotic drive system with the property that males heterozygous for an SD-bearing chromosome 2 and an SD⁺-bearing homolog transmit the SD-bearing chromosome almost exclusively. This distorted segregation is the consequence of an induced dysfunction of those sperm that receive the SD⁺ homolog. From previous studies, two loci have been implicated in this phenomenon: the Sd locus which is required to produce distortion, and the Responder (Rsp) locus that is the site at which Sd acts. There are two allelic alternatives of Rsp—sensitive (Rsp^sens) and insensitive (Rsp^ins); a chromosome carrying Rsp^ins is not distorted by SD. In the present study, the function and location of each of these elements was examined by a genetic and cytological characterization of X-ray-induced mutations at each locus. The results indicate the following: (1) the Rsp locus is located in the proximal heterochromatin of 2R; (2) a deletion for the Rsp locus renders a chromosome insensitive to distortion; (3) the Sd locus is located to the left of pr (2-54.5), in the region from 37D2-D7 to 38A6-B2 of the salivary chromosome map; (4) an SD chromosome deleted for Sd loses its ability to distort; (5) there is another important component of the SD system, E(SD), in or near the proximal heterochromatin of 2L, that behaves as a strong enhancer of distortion. The results of these studies allow a reinterpretation of results from earlier analyses of the SD system and serve to limit the possible mechanisms to account for segregation distortion.     

The Entire Compound Autosomes of DROSOPHILA MELANOGASTER

Three new unusual compound chromosomes have been synthesized in Drosophila melanogaster. They consist of two homologous autosomes joined together in the new order: right arm, left arm, centromere, left arm, right arm, for each of the two major autosomes, and one in which chromosomes 2 and 3 have been combined in the order: right arm of 2, left arm of 2, centromere, left arm of 3, right arm of 3. The attachments of the autosomal arms were accomplished by obtaining chromosome breaks at or very close to the ends of the left arms of the autosomes such that no essential chromosome material has been removed; the compounds derived from them are therefore referred to as entire compounds. These large chromosomes are recovered in progeny with frequencies lower than expectation partly because of zygote mortality associated with these chromosomes, and partly because of a failure of spermiogenesis.     

Deficiency Analysis of the Tip of Chromosome 3R in DROSOPHILA MELANOGASTER

Region 98EF-100F in chromosome 3 is interesting for genetic analysis because it contains a number of genes of developmental importance. Although there are no preexisting simple deficiency stocks, this region is amenable to genetic manipulation using other types of rearrangements. In the present investigation we obtained deficiencies by combining the terminal deficiencies formed by segregation of Y;3 translocations with a series of duplications of the tip of 3R, both from Y;3 translocations with different breakpoints and from 3;1 duplications in which the 3R tip is carried as a second arm on the X chromosome. Analysis of such synthetic deficiencies reveals five haplo-abnormal loci in the 98A-100F interval. These include a haplolethal site, a newly described Minute and three previously reported Minute mutations. The newly discovered Minute has been designated M(3)99D and is localized cytologically to bands 99D1-9. The three previously reported Minute loci in the region have been localized more precisely: M(3)1 to bands 99B5-9, M(3)f to bands 99E4-F1 and M(3)g to region 100C-F. In addition, we have been able to obtain synthetic deficiencies uncovering all of the intervals from 99B5 to 100B. These deficiencies will be useful for future genetic and molecular analyses of the genes that map within the right tip of chromosome 3.     

Genetic Analysis of the Centromeric Heterochromatin of Chromosome 2 of DROSOPHILA MELANOGASTER: Deficiency Mapping of Ems-Induced Lethal Complementation Groups

Until recently, little was known of the genetic constitution of the heterochromatic segments of the major autosomes of Drosophila melanogaster . Our previous report described the genetic dissection of the proximal, heterochromatic region of chromosome 2 of Drosophila melanogaster by means of a series of overlapping deficiencies generated by the detachment of compound second autosomes (Hilliker and Holm 1975). Analysis of these deficiencies by inter se complementation, pseudo-dominance tests with proximal mutations and allelism tests with known deficiencies provided evidence for the existence of at least two loci between the centromere and the light locus in 2L and one locus in 2R between the rolled locus and the centromere. These data in conjunction with cytological observations demonstrated that light and rolled and three loci lying between them are located within the proximal heterochromatin of the second chromosome.——The present report describes the further analysis of this region through the induction with ethyl methanesulphonate (EMS) of recessive lethals allelic to the 2L and 2R proximal deficiencies associated with the detachment products. Analysis of the 118 EMS-induced recessive lethals and visible mutations recovered provided evidence for seven loci in the 2L heterochromatin and six loci in the 2R heterochromatin, with multiple alleles being obtained for most sites. Of these loci, one in 2L and two in 2R fall near the heterochromatic-euchromatic junctions of 2L and 2R respectively. None of the 113 EMS lethals behaved as a deficiency, implying that the heterochromatic loci uncovered in this study represent nonrepetitive cistrons. Thus functional genetic loci are found in heterochromatin, albeit at a very low density relative to euchromatin.     

Genetics of Catalase in DROSOPHILA MELANOGASTER: Rates of Synthesis and Degradation of the Enzyme in Flies Aneuploid and Euploid for the Structural Gene

A screen for allelic variants of the enzyme catalase indicated that the Cat⁺ locus is essentially monomorphic in D. melanogaster. Segmental aneuploidy was used to screen the genome for a dosage-sensitive region for catalase activity. One region, 75D–78A on the polytene chromosome map of 3L, exhibited a hyperploid/euploid ratio of enzyme activity of 1.5. Further dissection localized the region to 75D–76A. We suggest that this region contains the structural locus for catalase in D. melanogaster.

Simple methods have been developed using the specific inhibitor, 3-amino-1,2,4-triazole, for the direct analysis of rates of synthesis and degradation of the Cat⁺ gene product. Based on kinetic studies of catalase synthesis in flies aneuploid and euploid for region 75D–76B, we suggest that these techniques can be readily applied to an examination of mutants that control the expression of the structural gene for catalase in Drosophila.

     

A Comprehensive Study of Genic Variation in Natural Populations of Drosophila melanogaster. III. Variations in Genetic Structure and Their Causes between Drosophila melanogaster and Its Sibling Species Drosophila simulans

The natural populations of Drosophila melanogaster and Drosophila simulans were compared for their genetic structure. A total of 114 gene-protein loci were studied in four mainland (from Europe and Africa) and an island (Seychelle) populations of D. simulans and the results were compared with those obtained on the same set of homologous loci in fifteen worldwide populations of D. melanogaster. The main results are as follows: (1) D. melanogaster shows a significantly higher proportion of loci polymorphic than D. simulans (52% vs. 39%, P<0.05), (2) both species have similar mean heterozygosity and mean number of alleles per locus, (3) the two species share some highly polymorphic loci but they do not share loci that show high geographic differentiation, and (4) D. simulans shows significantly less geographic differentiation than D. melanogaster. The differences in genetic differentiation between the two species are limited to loci located on the X and second chromosomes only; loci on the third chromosome show similar level of geographic differentiation in both species. These two species have previously been shown to differ in their pattern of variation for chromosomal polymorphisms, quantitative and physiological characters, two-dimensional electrophoretic (2DE) proteins, middle repetitive DNA and mitochondrial DNA. Variation in niche-widths and/or genetic "strategies" of adaptation appear to be the main causes of differences in the genetic structure of these two species.     

Post-Translational Modification of Xanthine Dehydrogenase in a Natural Population of DROSOPHILA MELANOGASTER

Second chromosomes of D. melanogaster were isolated from a single natural population, and 40 were analyzed by gel-sieving electrophoresis for the presence of polymorphic loci on chromosome 2 that act to modify xanthine dehydrogenase and/or aldehyde oxidase, whose structural genes map to chromosome 3. Clear evidence of polymorphism for one or more xanthine dehydrogenase modifier loci was obtained.     

Selection for Male Recombination in DROSOPHILA MELANOGASTER

Two-way selection for male recombination over seven intervals of the third chromosome in Drosophila melanogaster was practiced for nine generations followed by relaxed selection for five generations. Significant responses in both directions were observed but these mainly occurred in early generations in the low line and in later generations in the high line. Divergence of male recombination frequencies between the two selection lines was not restricted to any specific region but occurred in every measured interval of the chromosome. However, right-arm intervals showed a more pronounced response than either left-arm intervals or the centromeric region. Correlated responses in sterility and distortion of transmission ratios occurred as a result of selection for male recombination. Cluster distributions of male recombinants suggested a mixture of meiotic and late gonial events but relative map distances more closely resembled those of the salivary chromosome than standard meiotic or mitotic distances. Patterns of male recombination over time in both second and third chromosomes strongly suggested a major effect associated with the presence of third chromosomes from the Harwich strain. Evidence was also found for modifiers with relatively small effects located in other regions of the genome. The overall results are interpreted in terms of a two-component model of hybrid dysgenesis.     

Molecular Genetics of the ROSY-ACE Region of DROSOPHILA MELANOGASTER

Three hundred and fifteen kilobases of DNA from the rosy-Ace region on chromosome 3R of D. melanogaster have previously been cloned and extensively characterized. We describe the isolation of nine new deficiency mutants that break within the 315-kb interval. The position of these breakpoints on the DNA map was determined by in situ and Southern hybridization. Further, we more precisely mapped the breakpoints of several deletions previously analyzed. The results permit us to delimit sequences essential to the known complementation groups in the region within approximately 20 kb in most cases. However, one gene, B16-1, is shown to contain essential sequences that span about 50 kb. Also, we demonstrate by overlapping deficiencies that a 45-kb DNA segment from the region, which includes one known complementation group, allows limited survival when deleted.     

Glutamate oxalate transaminase (GOT) genetics in Mus musculus: Linkage,polymorphism, and phenotypes of the Got-2 and Got-1 loci

A comparison of electrophoretic patterns of F₁ and backcross progeny of two inbred strains of mice has revealed a new autosomal variant of the mitochondrial form of GOT. The loci controlling the production of the soluble and mitochondrial forms of GOT have been designated Got-1 and Got-2, respectively. The two alleles of the Got-2 locus have been designated Got-2 ^a and Got-2 ^b, which represent the slow- and fast-migrating electrophoretic forms. Twenty-seven inbred strains of mice have been classified for Got-2 ^a and Got-2 ^b. It has been demonstrated that the polymorphism of Got-2 is widely distributed in feral mice. Got-2 was shown to be linked to Es-1, and evidence is also presented for linkage between Got-2 and Es-2, Es-5, and oligosyndactyly (Os). The absence of linkage of Got-2 to seven other loci has also been demonstrated. GOT was expressed in vitro in cell lines derived from human and mouse tissues.     

Genic Heterogeneity at Two Alcohol Dehydrogenase Loci in DROSOPHILA PSEUDOOBSCURA and DROSOPHILA PERSIMILIS

A sequential electrophoretic survey of the second chromosome loci, alcohol dehydrogenase-6 (Adh-6) and octanol dehydrogenase ( Odh), was performed on 147 isochromosomal lines of Drosophila pseudoobscura and 60 lines of its sibling species, D. persimilis. Gels run with a variety of acrylamide concentrations and buffer pH's revealed the presence of 18 alleles of Adh-6 in the two species, where only eight had been previously detected by conventional electrophoretic methods. Only two alleles were added with our techniques to the previous total of nine in both species at the largely monomorphic Odh locus. Both enzymes show a predominance of one allele, with the other variants being fairly rare. There was no evidence of increased genetic divergence between the two species, but we found a striking increase in differentiation of Adh-6 alleles between the main body of D. pseudoobscura populations and the conspecific isolate from Bogotá, Colombia. These results are compared with our previous surveys of xanthine dehydrogenase in these species and discussed in reference to theories of genic polymorphism.     

Genetics of esterases in Drosophila. III. Influence of different chromosomes on esterase pattern in Drosophila

The present report presents the results of starch and polyacrylamide gel electrophoretic studies of the influence of the X chromosome on the expression of esterase-6 in D. melanogaster × D. simulans hybrids heterozygous for locus Est-6 as well as studies of the influence of autosomes on esterase expression in Drosophila of the virilis group. A differential expression of esterase-6 has been detected in D. melanogaster × D. simulans hybrid males. A differential decrease in the activity of esterase-6 (both F and S allozymes) derived from D. melanogaster has been noted. In hybrid females, the activity of parental esterases is the same. It is suggested that the X chromosome regulates the expression of esterase-6 in D. melanogaster. Analysis of individuals obtained in different schemes of crosses between different species of Drosophila of the virilis group by use of stocks marked with mutations in various chromosomes indicates that other autosomes (in particular, autosomes 4 and 5) also influence the phenotypic expression of esterases (which are controlled by genes located on the second chromosome).     

The Genetics of a Small Autosomal Region of DROSOPHILA MELANOGASTER Containing the Structural Gene for Alcohol Dehydrogenase. VI. Induced Revertants of Scutoid

Twenty-six induced revertants of Scutoid (Sco), a dominant mutation of Drosophila melanogaster, have been characterized genetically. Sco is an unusual mutation, involving two small reciprocal transpositions within the region 35A4 to 35C5 of chromosome arm 2L. One of these transpositions juxtaposes the noc and l(2)br28 loci. We suggested previously that the Sco phenotype results from the "fusion" of noc and l(2)br28. In support of this idea we now show that 23 of 26 revertants of Sco are noc^-, indeed the majority are either chromosome aberrations broken between noc and l(2)br28 or deletions of these loci from the mutant chromosome. However, some revertants of Sco are rather more complex, and their properties suggest an interaction between the pu-noc and l(2)br28-l(2)br37 regions of chromosome arm 2L and also demonstrate the genetic complexity of the el-noc region.     

FISH analysis of<Emphasis Type="Italic"> Drosophila melanogaster</Emphasis> heterochromatin using BACs and<Emphasis Type="Italic"> P</Emphasis> elements

The heterochromatin of chromosomes 2 and 3 of Drosophila melanogaster contains about 30 essential genes defined by genetic analysis. In the last decade only a few of these genes have been molecularly characterized and found to correspond to protein-coding genes involved in important cellular functions. Moreover, several predicted genes have been identified by annotation of genomic sequence that are associated with polytene chromosome divisions 40, 41 and 80 but their locations on the cytogenetic map of the heterochromatin are still uncertain. To expand our current knowledge of the genetic functions located in heterochromatin, we have performed fluorescence in situ hybridization (FISH) mapping to mitotic chromosomes of nine bacterial artificial chromosomes (BACs) carrying several predicted genes and of 13 P element insertions assigned to the proximal regions of 2R and 3L. We found that 22 predicted genes map to the h46 region of 2R and eight map to the h47 regions of 3L. This amounts to at least 30 predicted genes located in these heterochromatic regions, whereas previous studies detected only seven vital genes. Finally, another 58 genes localize either in the euchromatin-heterochromatin transition regions or in the proximal euchromatin of 2R and 3L. Edited by: B. McKeeN. Corradini and F. Rossi contributed equally to this work     

Cytogenetic Mapping of Enzyme Loci on Chromosomes J and U of DROSOPHILA SUBOBSCURA

Enzyme loci located on chromosome J and U were mapped cytologically by means of a Y translocation technique. A linkage map of the two chromosomes was established in a parallel experiment and the recombination frequency in different regions of the chromosomes determined. A comparison of the cytogenetic localization of the enzyme genes in D. subobscura and D. melanogaster indicates that many paracentric inversions must have taken place in the course of divergent evolution. However, no displacements of genes from one element to another due to pericentric inversions, reciprocal translocations or transposing elements can be observed. In spite of the large number of structural rearrangements that have occurred in the phylogeny of the genus Drosophila, gross similarities of banding pattern in homologous regions of the chromosomes of the two species become apparent.     

Orientation Disruptor (ord): A Recombination-Defective and Disjunction-Defective Meiotic Mutant in DROSOPHILA MELANOGASTER

The effects of a semidominant autosomal meiotic mutant, orientation disruptor (symbol: ord), located at 2–103.5 on the genetic map and in region 59B-D of the salivary map, have been examined genetically and cytologically. The results are as follows. (1) Crossing over in homozygous females is reduced to about seven percent of controls on all chromosomes, with the reduction greatest in distal regions. (2) Crossing over on different chromosomes is independent. (3) Reductional nondisjunction of any given chromosome is increased to about thirty percent of gametes from homozygous females. The probability of such nondisjunction is the same among exchange and nonexchange tetrads with the exception that a very proximal exchange tends to regularize segregation. (4) Equational nondisjunction of each chromosome is increased to about ten percent of gametes in homozygous females; this nondisjunction is independent of exchange. (5) The distributive pairing system is operative in homozygous females. (6) In homozygous males, reductional nondisjunction of each chromosome is increased to about ten percent, and equational nondisjunction to about twenty percent, of all gametes. (7) Cytologically, two distinct meiotic divisions occur in spermatocytes of homozygous males. The first division looks normal although occasional univalents are present at prophase I and a few lagging chromosomes are seen at anaphase I. However, sister chromatids of most chromosomes have precociously separated by metaphase II. Possible functions of the ord⁺ gene are considered.     

Genetic Control of Adh Expression in DROSOPHILA MELANOGASTER

Natural variants displaying different levels of expression of the gene for alcohol dehydrogenase (Adh) were subjected to genetic mapping experiments. The strains studied carry one of the two common electrophoretic forms of the enzyme. The difference among Adh-fast strains appears to be due to multiple loci with trans-acting effects. Differences among Adh-slow strains are due to modifiers or quantitative sites located very close to the structural gene (less than 0.05 map unit) or part of it. The modifiers detected in the Adh^s strains seem to operate only on the structural allele in the cis-position.—A modifier that affects the ratio of ADH levels in larvae and adults was also detected in the Adh^s strains. This modifier is also closely linked to Adh and is cis-acting.     

Genetic control and intracellular localization of glutamate oxaloacetic transaminase in maize

Glutamate oxaloacetic transaminase (L-aspertate: 2-oxoglutarate aminotransferase, E.C. 2.6.1.1; GOT) was found to occur in five distinct electrophoretic forms in different tissue extracts from a number of highly inbred strains of Zea mays L. No major qualitative differences were detected in the various tissues examined, and the isozyme patterns did not undergo changes during temporal development of any given inbred strain. Cell fractionation studies showed one isozyme to be associated with the mitochondria (mGOT), another to be exclusively associated with the soluble fraction (sGOT), and a third to be associated with the glyoxysomes (gGOT). The glyoxysomal form occurs as two electrophoretically distinct variants which exist in different inbred strains of maize. The gGOT variants are under the control of two codiminant alleles (Got1A and Got1B) at the Got1 locus (isozyme5, gGOT). The genetic data and gene dosage effects suggest that GOT in maize is functionally a dimer.     

Third-Chromosome Mutagen-Sensitive Mutants of DROSOPHILA MELANOGASTER

A total of 34 third chromosomes of Drosophila melanogaster that render homozygous larvae hypersensitive to killing by chemical mutagens have been isolated. Genetic analyses have placed responsible mutations in more than eleven complementation groups. Mutants in three complementation groups are strongly sensitive to methyl methanesulfonate, those in one are sensitive to nitrogen mustard, and mutants in six groups are hypersensitive to both mutagens. Eight of the ten loci mapped fall within 15% of the genetic map that encompasses the centromere of chromosome 3. Mutants from four of the complementation groups are associated with moderate to strong meiotic effects in females. Preliminary biochemical analyses have implicated seven of these loci in DNA metabolism.