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R J MacIntyre

Publications and source records attributed to R J MacIntyre.

At least 19 recordsLinked to original sources
PubMed2001

Genetic and molecular characterization of a variegating hsp70-acZ fusion gene in the euchromatic 31 B region of Drosophila melanogaster.

A hsp70-lacZ fusion gene introduced into Drosophila melanogaster at the euchromatic 31B region by Pelement transformation displayed a variegated expression with respect to the lacZ fusion protein in the salivary gland cells under heat-shock conditions. The variegation is also reflected by the chromosome puffing pattern. Subsequent transposition of the 31B P element to other euchromatic positions restored wild-type activity, that is, a nonvariegated phenotype. A lower developmental temperature reduced the amount of expression under heat-shock conditions, similar to genes undergoing position-effect variegation (PEV). However, other modifiers of PEV did not affect the expression pattern of the gene. These results show a novel euchromatic tissue-specific variegation that is not associated with classical heterochromatic PEV.

Animals↗
PubMed1997

Molecular cloning of a family of xenobiotic-inducible drosophilid cytochrome p450s: evidence for involvement in host-plant allelochemical resistance.

Cytochrome P450s constitute a superfamily of genes encoding mostly microsomal hemoproteins that play a dominant role in the metabolism of a wide variety of both endogenous and foreign compounds. In insects, xenobiotic metabolism (i.e., metabolism of insecticides and toxic natural plant compounds) is known to involve members of the CYP6 family of cytochrome P450s. Use of a 3' RACE (rapid amplification of cDNA ends) strategy with a degenerate primer based on the conserved cytochrome P450 heme-binding decapeptide loop resulted in the amplification of four cDNA sequences representing another family of cytochrome P450 genes (CYP28) from two species of isoquinoline alkaloid-resistant Drosophila and the cosmopolitan species Drosophila hydei. The CYP28 family forms a monophyletic clade with strong regional homologies to the vertebrate CYP3 family and the insect CYP6 family (both of which are involved in xenobiotic metabolism) and to the insect CYP9 family (of unknown function). Induction of mRNA levels for three of the CYP28 cytochrome P450s by toxic host-plant allelochemicals (up to 11.5-fold) and phenobarbital (up to 49-fold) corroborates previous in vitro metabolism studies and suggests a potentially important role for the CYP28 family in determining patterns of insect-host-plant relationships through xenobiotic detoxification.

Amino Acid Sequence↗
PubMed1994

A model of the Drosophila melanogaster glycerol-3-phosphate oxidase-encoding gene.

We have sequenced the Drosophila melanogaster gene encoding the mitochondrial (mt) enzyme glycerol-3-phosphate oxidase (GPO; EC 1.1.99.5) that is involved in flight and alcohol metabolism. Available data suggests a simple model for this gene that includes four exons. Exon I contains a mt import signal, exon II, a transmembrane segment and an FAD-binding site, and exon IV, an iron-sulfur centre.

Alternative Splicing↗
PubMed1994

Molecular evolution: codes, clocks, genes and genomes.

The discoveries, advancements and continuing controversies in the field of molecular evolution are reviewed. Topics summarized are (1) the evolution of the genetic code, (2) gene evolution including the demonstration of homology, estimation of sequence divergence, phylogenetic trees, the molecular clock and the origin of genes and gene families by various genetic mechanisms, and (3) eukaryotic genome evolution, including the highly repeated satellite sequences, the interspersed and potentially mobile repeated sequences and the unique sequence fraction of the genome.

Biological Evolution↗
PubMed1991

Molecular cloning and transformation of cyclodiene resistance in Drosophila: an invertebrate gamma-aminobutyric acid subtype A receptor locus.

Cyclodiene resistance represents 60% of the reported cases of insecticide resistance and is also present in vertebrates. Resistance is due to insensitivity of the cyclodiene/picrotoxinin binding site on the gamma-aminobutyric acid subtype A (GABAA) receptor-chloride ionophore complex. Following isolation of cyclodiene-resistant Drosophila mutants, we report the cloning of the locus conferring resistance via a "chromosomal walk" and rescue of the susceptible phenotype by P-element-mediated germ-line transformation. Amino acid sequence analysis of a cDNA from the locus reveals homology with vertebrate GABAA subunits. To our knowledge, this represents the first cloning of an invertebrate GABA receptor and also allows us to manipulate the resistance status of an insect via germ-line transformation. This gene may be useful as a selectable marker in other insect systems.

Amino Acid Sequence↗
PubMed1991

Molecular cloning and analysis of the chromosomal region 26A of Drosophila melanogaster.

Region 26A of the second chromosome of Drosophila melanogaster has been extensively characterized at the genetic level. We report here the cloning of virtually the entire 26A region via a bidirectional chromosome walk. Deletion and translocation breakpoints in the 26A interval have been localized at the molecular level by both chromosomal in situ hybridization and Southern analysis. The locations of the genetically defined loci in this chromosomal region have also been correlated with transcriptional units mapped onto the DNA of the proximal region of the chromosomal walk. The position of the alpha-glycerophosphate dehydrogenase (alpha-Gpdh) gene in 26A5-7 has been confirmed and a putative transcriptional unit for the beta-galactosidase-1 (beta-Gal-1) gene has been identified in the 26A7-9 interval.

Animals↗
PubMed1990

Cryopreservation of Drosophila melanogaster embryos.

There is an urgent need to preserve the ever-increasing number (greater than 30,000) of different genetic strains of D. melanogaster that are maintained in national and international stock centres and in the laboratories of individual investigators. In all cases, the stocks are maintained as adult populations and require transfer to fresh medium every two to four weeks. This is not only costly in terms of materials, labour and space, but unique strains are vulnerable to accidental loss, contamination, and changes in genotype that can occur during continuous culture through mutation, genetic drift or selection. Although cryopreservation of Drosophila germ-plasm would be an enormous advantage, many attempts using conventional procedures have been unsuccessful. D. melanogaster embryos are refractory to conventional cryopreservation procedures because of the contravening conditions required to minimize mortality resulting from both intracellular ice formation and chilling injury at subzero temperatures. To overcome these obstacles, we have developed a vitrification procedure that precludes intracellular ice formation so that the embryos can be cooled and warmed at ultra-rapid rates to minimize chilling injury, and have recovered viable embryos following storage in liquid nitrogen. In a series of 53 experiments, a total of 3,711 larvae emerged from 17,280 eggs that were cooled in liquid nitrogen (18.4 +/- 8.8%). Further, using a subset from this population, approximately 3% of the surviving larvae (24/800) developed into adults. These adults were fertile and produced an F1 generation.

Animals↗
PubMed1990

The isolation of the acid phosphatase-1 gene of Drosophila melanogaster and a chromosomal breakpoint inducing its position effect variegation.

Over 120 kb of contiguous genomic DNA sequence derived from the 99C-99D region of the Drosophila melanogaster third chromosome were isolated by molecular cloning. Sequences within this region required for the expression of the lysosomal gene-enzyme system acid phosphatase-1 (Acph-1) were identified by both P element-mediated germline transformation and transient expression and lie within a single 5 kb fragment. Acph-1 is encoded by a 2.1 kb poly(A)+ RNA transcript, which is expressed throughout development. Enzyme activity peaks also correlate with increases in RNA abundance. The ca-74 deletion, which exhibits position effect variegation at the Acph-1 gene (Frisardi and MacIntyre 1984), was also partially characterized. The variegating ca-74 breakpoint is located approximately 20 kb proximal to the Acph-1 gene. Results suggest that the heterochromatin at this breakpoint comprises highly repetitive or satellite DNA.

Acid Phosphatase↗
PubMed1989

A genetic and molecular analysis of P-induced mutations at the glucose-6-phosphate dehydrogenase locus in Drosophila melanogaster.

We examined P factor induced mutations of the Zw gene of Drosophila melanogaster in order to learn more about the site specificity of such mutations. Approximately 70,000 chromosomes were screened using a powerful positive selection scheme. As only two mutants were discovered, Zw is a "cold spot" for transposable element insertion. One mutation involved a complex P element associated chromosomal rearrangement which was used to define the orientation of the gene with respect to the centromere of the X chromosome. The second mutation was either a simple, non-dysgenically induced point mutation or a very unstable insertion.

Animals↗
PubMed1989

Structural characterization of the alpha-glycerol-3-phosphate dehydrogenase-encoding gene of Drosophila melanogaster.

In Drosophila, multiple isoforms of alpha-glycerol-3-phosphate dehydrogenase (sn-glycerol-3-phosphate: NAD+ 2-oxidoreductase, EC 1.1.1.8) are produced in a tissue- and stage-specific manner. To understand the underlying molecular basis of these isoforms, we have sequenced a 5.8-kilobase region of the Drosophila genome that contains the entire Gpdh locus. Primer-extension and RNase protection assays show that the gene consists of eight exons and has a single transcription-start point. RNase protection mapping and comparison of the genomic sequence from three different cDNA clones reveal that three protein isoforms of glycerol-3-phosphate dehydrogenase are produced by alternative processing of 3' exons. Two of the isoforms differ from the third by the addition of either three or ten amino acids to their C-terminal ends. Transcripts corresponding to two of the isoforms are expressed during both larval and adult stages, while the third isoform is produced only in adults.

Amino Acid Sequence↗
PubMed1988

A genetic analysis of the alpha-glycerophosphate oxidase locus in Drosophila melanogaster.

The gene for alpha-glycerophosphate oxidase, the nuclear encoded mitochondrial enzyme of the alpha-glycerophosphate cycle (alpha GP); has been mapped in Drosophila melanogaster. Several interstitial deficiencies in region 50c-53AB of chromosome 2R were used to localize the structural gene to 52D2-5. In addition, mutations of alpha GPO were generated; alpha GPO mutants are viable yet flightless. Interactions of alpha GPO with alpha-glycerophosphate dehydrogenase (alpha GPDH), the cytoplasmic enzyme of the alpha GP cycle, were investigated through the synthesis of a series of alpha GPDHnull-alpha GPOnull double mutants. Of the six double null mutants constructed, four alpha GPDH-alpha GPO double nulls are viable and flightless. Two double mutants, however, exhibit an allelic-dependent synthetic lethal phenotype.

Aneuploidy↗
PubMed1987

Drosophila melanogaster homologs of the raf oncogene.

A murine v-raf probe, representing the kinase domain, was used to identify two unique loci in Drosophila melanogaster DNA. The most closely related to v-raf was mapped by in situ hybridization to position 2F5-6 (Draf-1) on the X chromosome, whereas the other raf-related gene (Draf-2) was found at position 43A2-5 on chromosome 2. The nucleotide and amino acid homologies of Draf-1 to the kinase domain of v-raf are 61 and 65%, respectively. The large amount of a 3.2-kilobase Draf-1 transcript detected in eggs as a maternal message decreases during embryonic development, and significant steady-state levels are observed throughout the remainder of morphogenesis. We speculate that the Draf-1 locus plays an important role in early embryogenesis.

Amino Acid Sequence↗
PubMed1986

Isolation of a genomic clone for Drosophila sn-glycerol-3-phosphate dehydrogenase using synthetic oligonucleotides.

A genomic clone containing Drosophila sn-glycerol-3-phosphate dehydrogenase sequences has been isolated using a mixture of synthetic tridecanucleotides as a hybridization probe. The clone as well as the synthetic probe mixture was found to hybridize to an abundant poly(A)+ RNA of 1700 bases. A partial DNA sequence obtained for a 40-amino acid region containing the oligonucleotide hybridization site was found to agree with the known Drosophila protein sequence data for this region of the protein. In situ hybridization of this clone to the polytene chromosomes of wild type flies and flies bearing chromosomal aberrations that delimit the Gpdh+ locus have allowed us to decisively place the gene in the distal region of 26A on the left arm of the second chromosome.

Amino Acid Sequence↗
PubMed1984

Cytogenic mapping and isolation of mutations of the beta-Gal-1 locus of Drosophila melanogaster.

The structural gene beta-Gal-1 encoding a beta-galactosidase (EC 3.2.1.23) of Drosophila melanogaster has been mapped by two independent genetic approaches. In the first, gene dosage dependent variation in beta-galactosidase activity levels in segmental aneuploids, generated from crosses of Y-autosome translocation stocks, was determined quantitatively. A dosage sensitive region on the left arm of chromosome 2 was identified and mapped to region 26A7-9. In the second approach, two null activity variants were isolated from wild populations. It was shown by deletion analysis that these nulls map to the same region as that determined by the segmental aneuploidy method. The results of an EMS mutagenesis screen showed that, besides the beta-Gal-1 locus, there are four loci defined by recessive lethal mutations which map in the 26A7-9 region.

Animals↗
PubMed1984

Position effect variegation of an acid phosphatase gene in Drosophila melanogaster.

X-ray mutagenesis has produced a series of deficiencies in a duplication of part of the third chromosome containing the acid phosphatase gene (Acph-1) in Drosophila melanogaster. In one of these deficiencies, Acph-1 is shown to be undergoing position effect variegation. Naturally occurring electrophoretic variants of the enzyme were used to visualize and determine quantitatively the extent of variegation of the allele which is cis to the heterochromatic breakpoint. Alteration of genotypic background and temperature provided further evidence for position effect. Rocket immunoelectrophoresis was used to correlate the levels of acid phosphatase activity and protein in flies containing the deficiency. A novel result indicates that the variegation is not the consequence of an averaging of active and inactive cells, but rather due to a quantitative alteration of gene activity within at least some individual cells.

Acid Phosphatase↗
PubMed1983

Biphasic expression and function of glucose dehydrogenase in Drosophila melanogaster.

Glucose dehydrogenase (GO) was found to be expressed during the pupal stage in both sexes of Drosophila melanogaster, but is limited to the male ejaculatory duct at the adult stage. During copulation GO is transferred from males to females. Mutational analysis of the Go locus indicates that a single structural gene encodes the pupal and ejaculatory duct GO. Thus an example of an enzyme structural gene switching from non-sex-limited to sex-limited expression has been found. Go mutants are recessive lethals exhibiting a late pupal effective lethal phase. These mutants can be rescued by excising the anterior end of the pupal case 0-2 days prior to the normal adult emergence time. It appears that the function of GO in pupae is to aid in the degradation of the puparium cuticle in preparation for the eclosion of the adult.

Animals↗
PubMed1983

A cytogenetic analysis of the chromosomal region surrounding the alpha-glycerophosphate dehydrogenase locus of Drosophila melanogaster.

The chromosomal region surrounding the structural gene for alpha-glycerophosphate dehydrogenase (alpha Gpdh, 2-20.5) of Drosophila melanogaster has been studied in detail. Forty-three EMS-induced recessive lethal mutations and five previously identified visible mutations have been localized within the 25A-27D region of chromosome 2 by deficiency mapping and in some cases by a recombination analysis. The 43 lethal mutations specify 17 lethal loci. alpha Gpdh has been localized to a single polytene chromosome band, 25F5, and there apparently are no lethals that map to the alpha Gpdh locus.

Animals↗