scholarly journals Molecular control of the induction of alcohol dehydrogenase by ethanol in Drosophila melanogaster larvae.

Genetics ◽  
1990 ◽  
Vol 124 (4) ◽  
pp. 881-888 ◽  
Author(s):  
A M Kapoun ◽  
B W Geer ◽  
P W Heinstra ◽  
V Corbin ◽  
S W McKechnie
Keyword(s):  
Drosophila Melanogaster ◽  
Alcohol Dehydrogenase ◽  
Dna Sequence ◽  
P Element ◽  
Start Site ◽  
Major Pathway ◽  
Molecular Control ◽  
Adh Gene ◽  
Regulatory Dna ◽  
Transcript Start Site

Abstract The activity of alcohol dehydrogenase (ADH:EC 1.1.1.1), the initial enzyme in the major pathway for ethanol degradation, is induced in Drosophila melanogaster larvae by low concentrations of dietary ethanol. Two lines of evidence indicate that the metabolic products of the ADH pathway for ethanol degradation are not directly involved in the induction of Adh. First, the accumulation of the proximal transcript in Adhn2 larvae was increased when the intracellular level of ethanol was elevated. In addition, the ADH activity, the proximal Adh mRNA, and the intracellular concentration of ethanol were elevated coordinately in wild-type larvae fed hexadeuterated-ethanol, which is metabolized more slowly than normal ethanol. An examination of P element transformant lines with specific deletions in the 5' regulatory DNA of the Adh gene showed that a DNA sequence between +527 and +604 of the distal transcript start site is essential for the induction of the Adh gene [corrected]. The DNA sequence between -660 and about -5000 of the distal transcript start site was important for the down-regulation of the induction response.

scholarly journals Effects of a transposable element insertion on alcohol dehydrogenase expression in Drosophila melanogaster.

Genetics ◽  
1995 ◽  
Vol 140 (2) ◽  
pp. 667-677 ◽  
Author(s):  
R C Dunn ◽  
C C Laurie
Keyword(s):  
Drosophila Melanogaster ◽  
Alcohol Dehydrogenase ◽  
Transposable Element ◽  
Dna Sequence ◽  
Natural Populations ◽  
P Element ◽  
Dna Polymorphisms ◽  
Threefold Increase ◽  
Adh Gene ◽  
High Level

Abstract Variation in the DNA sequence and level of alcohol dehydrogenase (Adh) gene expression in Drosophila melanogaster have been studied to determine what types of DNA polymorphisms contribute to phenotypic variation in natural populations. The Adh gene, like many others, shows a high level of variability in both DNA sequence and quantitative level of expression. A number of transposable element insertions occur in the Adh region and one of these, a copia insertion in the 5' flanking region, is associated with unusually low Adh expression. To determine whether this insertion (called R142) causes the low expression level, the insertion was excised from the cloned R142 Adh gene and the effect was assessed by P-element transformation. Removal of this insertion causes a threefold increase in the level of ADH, clearly showing that it contributes to the naturally occurring variation in expression at this locus. Removal of all but one LTR also causes a threefold increase, indicating that the mechanism is not a simple sequence disruption. Furthermore, this copia insertion, which is located between the two Adh promoters and their upstream enhancer sequences, has differential effects on the levels of proximal and distal transcripts. Finally, a test for the possible modifying effects of two suppressor loci, su(wa) and su(f), on this insertional mutation was negative, in contrast to a previous report in the literature.

scholarly journals Identification of cis-regulatory elements required for larval expression of the Drosophila melanogaster alcohol dehydrogenase gene.

Genetics ◽  
1990 ◽  
Vol 124 (3) ◽  
pp. 637-646 ◽  
Author(s):  
V Corbin ◽  
T Maniatis
Keyword(s):  
Drosophila Melanogaster ◽  
Alcohol Dehydrogenase ◽  
Dna Sequences ◽  
Regulatory Elements ◽  
Malpighian Tubules ◽  
Proximal Promoter ◽  
Start Site ◽  
Wild Type ◽  
Tissue Specific ◽  
Adh Gene

Abstract The Alcohol dehydrogenase (Adh) genes of two distantly related species, Drosophila melanogaster and Drosophila mulleri, display similar, but not identical, patterns of tissue-specific expression in larvae and adults. The regulatory DNA sequences necessary for wild-type Adh expression in D. mulleri larvae were previously reported. In this paper we present an analysis of the DNA sequences necessary for wild-type Adh expression in D. melanogaster larvae. We show that transcription from the proximal promoter of the melanogaster Adh gene is regulated by a far upstream enhancer and two or more elements near the transcription start site. The enhancer is tissue specific and stimulates transcription to high levels in fat body and to lower levels in midgut and malpighian tubules whether linked to the proximal promoter or to a heterologous promoter. The enhancer activity localized to at least two discrete regions dispersed over more than 1.7 kb of DNA. Deletion of any one of these subregions reduces Adh transcription in all three larval tissues. Similarly, two regions immediately upstream of the proximal promoter start site are necessary for wild-type transcription levels in all three tissues. Thus, each of the identified regulatory elements is sufficient for low levels of Adh gene expression in all three larval tissues, but maximal levels of expression requires the entire set.

Rapid spread of a P element/Adh gene construct through experimental populations of Drosophila melanogaster

Genome ◽  
1993 ◽  
Vol 36 (6) ◽  
pp. 1169-1175 ◽  
Author(s):  
G. A. Meister ◽  
T. A. Grigliatti
Keyword(s):  
Drosophila Melanogaster ◽  
Alcohol Dehydrogenase ◽  
Transposable Elements ◽  
Transposable Element ◽  
Dehydrogenase Activity ◽  
P Element ◽  
P Elements ◽  
Alcohol Dehydrogenase Activity ◽  
Adh Gene ◽  
Experimental Populations

Transposable elements may be potential tools for the dispersal of engineered DNA through target insect populations. The utility of this hypothesis is predicated on the ability of transposable elements carrying a large DNA insert to rapidly disperse through a population. In addition, the inserted DNA must be replicated with a high degree of fidelity during this dispersal. We have monitored the ability of a transposable element with an inserted gene to spread through experimental populations and tested whether the passenger gene retains its ability to encode an active protein. Several Drosophila melanogaster laboratory populations were initiated with female flies that were null for alcohol dehydrogenase activity and contained no P elements. Most of the females were mated to males of the same strain; however, 1 or 10% of the females were mated to males from a strain that had previously been transformed with a helper P element and a P element/Adh gene construct. The dispersal of P elements to new genomes was monitored at each generation by randomly selecting females and performing DNA hybridization assays on dissected ovarian tissue. In addition, each female was tested for alcohol dehydrogenase activity using a simple histochemical assay. We find that, despite an approximate threefold increase in size, the P element constructs containing a functioning gene are still capable of rapid dispersal through the experimental populations. We also show that many of the inserted Adh genes still encode an active product.Key words: P element, transformation, Adh, transposable element.

Expression of an amylase - alcohol dehydrogenase chimeric gene in transgenic strains of Drosophila melanogaster

Genome ◽  
1993 ◽  
Vol 36 (5) ◽  
pp. 954-961 ◽  
Author(s):  
Allan A. Grunder ◽  
Ada Loverre-Chyurlia ◽  
Donal A. Hickey
Keyword(s):  
Drosophila Melanogaster ◽  
Alcohol Dehydrogenase ◽  
Dna Analysis ◽  
Glucose Repression ◽  
Chimeric Gene ◽  
P Element ◽  
Regulatory Sequences ◽  
Mrna Levels ◽  
Promoter Sequences ◽  
Adh Gene

A chimeric gene, consisting of 428 bp of the promoter sequences of the α-amylase gene of Drosophila melanogaster, fused to the transcribed region of the alcohol dehydrogenase (Adh) gene, was introduced into the genome of an Adhnull stock of Drosophila via P element mediated transformation. DNA analysis (Southern blotting) of three transformant strains confirmed the insertion of either one or two copies of the chimeric gene per strain. A histochemical study of ADH enzyme activity in dissected tissues of the transgenic larvae revealed that the chimeric Amy–Adh gene was expressed only in the posterior larval midgut and that this expression was repressed by dietary glucose, thus representing an expression pattern characteristic of the Amy gene. This indicates that the Amy upstream promoter sequences contain signals mediating both tissue specificity and glucose repression of the Adh structural gene in the transgenic larvae. The level of ADH activity expressed in transgenic flies was relatively low. This was paralleled by a low level of Adh mRNA, indicating a reduction in the transcriptional rate of the chimeric gene.Key words: Drosophila, germline transformation, chimeric gene, cis-regulatory sequences, α-amylase, alcohol dehydrogenase, tissue-specific expression, glucose repression, mRNA levels.

The ovarian, ecdysterone, and heat-shock-responsive promoters of the Drosophila melanogaster hsp27 gene react very differently to perturbations of DNA sequence

1987 ◽  
Vol 7 (3) ◽  
pp. 973-981
Author(s):  
E P Hoffman ◽  
S L Gerring ◽  
V G Corces
Keyword(s):  
Drosophila Melanogaster ◽  
Heat Shock ◽  
Dna Sequence ◽  
Germ Line ◽  
P Element ◽  
Position Effects ◽  
Consensus Sequences ◽  
Basal Expression ◽  
Upstream Promoter ◽  
Germ Line Transformation

The effect of various types of DNA sequence alterations on the activity of the ovarian, ecdysterone, and heat-shock-responsive promoters of the Drosophila melanogaster hsp27 gene was studied by P element-mediated germ line transformation. Regions of DNA required for proper expression of the gene under these different conditions were identified. Wild-type levels of transcription during oogenesis are dependent on two elements respectively located within a 64-base-pair (bp) fragment in the transcribed untranslated region and between -227 and -958 bp upstream of the transcription start site. This ovarian expression is particularly sensitive to both chromosomal position effects and an increased distance between the distal upstream promoter element and the TATAA homology. The ecdysterone-mediated expression during metamorphosis is dependent on a 145-bp domain including the TATAA box and additional upstream sequences that augment transcription by two- to five-fold. Finally, sequences necessary for heat shock expression are located much further upstream from hsp27 than those previously found for hsp70, although basal expression was correlated with the presence of more proximal heat shock consensus sequences.

scholarly journals Tissue-specific expression phenotypes of Hawaiian Drosophila Adh genes in Drosophila melanogaster transformants.

Genetics ◽  
1990 ◽  
Vol 125 (3) ◽  
pp. 599-610
Author(s):  
C Y Wu ◽  
J Mote ◽  
M D Brennan
Keyword(s):  
Drosophila Melanogaster ◽  
Tissue Distribution ◽  
Germ Line ◽  
P Element ◽  
Specific Expression ◽  
Position Effects ◽  
Tissue Specific ◽  
Hawaiian Drosophila ◽  
Adh Genes ◽  
Adh Gene

Abstract Interspecific differences in the tissue-specific patterns of expression displayed by the alcohol dehydrogenase (Adh) genes within the Hawaiian picture-winged Drosophila represent a rich source of evolutionary variation in gene regulation. Study of the cis-acting elements responsible for regulatory differences between Adh genes from various species is greatly facilitated by analyzing the behavior of the different Adh genes in a homogeneous background. Accordingly, the Adh gene from Drosophila grimshawi was introduced into the germ line of Drosophila melanogaster by means of P element-mediated transformation, and transformants carrying this gene were compared to transformants carrying the Adh genes from Drosophila affinidisjuncta and Drosophila hawaiiensis. The results indicate that the D. affinidisjuncta and D. grimshawi genes have relatively higher levels of expression and broader tissue distribution of expression than the D. hawaiiensis gene in larvae. All three genes are expressed at similar overall levels in adults, with differences in tissue distribution of enzyme activity corresponding to the pattern in the donor species. However, certain systematic differences between Adh gene expression in transformants and in the Hawaiian Drosophila are noted along with tissue-specific position effects in some cases. The implications of these findings for the understanding of evolved regulatory variation are discussed.

scholarly journals Molecular Control of the Induction of Alcohol Dehydrogenase by Ethanol in Drosophila Melanogaster Larvae

Genetics ◽  
1990 ◽  
Vol 126 (2) ◽  
pp. 477-477
Author(s):  
A M Kapoun ◽  
B W Geer ◽  
PWH Heinstra ◽  
V Corbin ◽  
S W McKechnie
Keyword(s):  
Drosophila Melanogaster ◽  
Alcohol Dehydrogenase ◽  
Molecular Control

scholarly journals Insertion of a copia element 5' to the Drosophila melanogaster alcohol dehydrogenase gene (adh) is associated with altered developmental and tissue-specific patterns of expression.

Genetics ◽  
1989 ◽  
Vol 121 (4) ◽  
pp. 787-794
Author(s):  
D J Strand ◽  
J F McDonald
Keyword(s):  
Drosophila Melanogaster ◽  
Alcohol Dehydrogenase ◽  
Transcriptional Start Site ◽  
Adult Life ◽  
Life Stages ◽  
Start Site ◽  
Wild Type ◽  
Dehydrogenase Gene ◽  
In Trans ◽  
Copia Element

Abstract The Drosophila melanogaster alcohol dehydrogenase gene (adh) is under the control of two separate promoters (proximal and distal) which are preferentially utilized at the larval and adult life stages, respectively. A variant alcohol dehydrogenase allele (RI-42) isolated from a natural population contains a copia retroviral-like transposable element inserted 240 bp upstream from the distal (adult) adh transcriptional start site. Levels of adh transcripts in the RI-42 variant are reduced in tissues and at life stages where copia is actively expressed and are affected in trans- by mutant alleles at the suppressor-of-white-apricot (su(wa] and suppressor-of-forked (su(f] loci. These suppressor genes have no effect on adh expression in wild-type Drosophila.

scholarly journals Analysis of sequences regulating larval expression of the Adh gene of Drosophila melanogaster.

Genetics ◽  
1991 ◽  
Vol 129 (3) ◽  
pp. 763-771
Author(s):  
N L Shen ◽  
E C Hotaling ◽  
G Subrahmanyam ◽  
P F Martin ◽  
W Sofer
Keyword(s):  
Enzyme Activity ◽  
Drosophila Melanogaster ◽  
Alcohol Dehydrogenase ◽  
Reference Gene ◽  
Base Pair ◽  
Regulatory Region ◽  
Regulatory Elements ◽  
Base Pair Deletion ◽  
Adh Gene ◽  
Tata Sequence

Abstract The effects of a series of eight, 50 base pair internal deletions in the 5' region upstream of the proximal transcription start site of the Adh gene of Drosophila melanogaster were examined in a quantitative assay. Mixtures of two plasmids, one bearing a deleted gene, the other with an intact reference gene, were injected into alcohol dehydrogenase-negative embryos. Third instar larvae of the injected generation were assayed for relative alcohol dehydrogenase enzyme activity. Quantitative analysis of the eight deletions indicated that two regions were required for any detectable enzyme activity and one region was required for appropriate tissue specificity. The remaining five deletions significantly decreased, but did not eliminate activity. When the deleted genes were placed on a plasmid with an intact reference gene, activities of all but one deletion were restored to levels equivalent to that of the intact reference gene (regardless of orientation). This restoration of activity did not occur when the regulatory region of the intact gene was replaced with the Hsp70 heat shock promoter nor when the 50-base pair deletion encompassed the region that includes the TATA sequence. The fact that seven of the eight deleted genes express activity in the presence of a reference gene on the same plasmid suggests that the deleted gene is controlled by regulatory elements in the reference gene. Further, these regulatory elements exhibit no preference for their own, more proximate, promoter.

scholarly journals Distribution and structure of cloned P elements from the Drosophila melanogaster P strain π2

1992 ◽  
Vol 60 (1) ◽  
pp. 33-41 ◽  
Author(s):  
Kevin O'Hare ◽  
Alan Driver ◽  
Stephen McGrath ◽  
Dena M. Johnson-Schiltz
Keyword(s):  
Drosophila Melanogaster ◽  
Transposable Elements ◽  
Dna Sequence ◽  
Genomic Library ◽  
Single Copy ◽  
P Element ◽  
Base Pairs ◽  
Single Base ◽  
P Elements ◽  
Element Sequence

SummaryP transposable elements of Drosophila melanogaster cloned from the strong P strain π2 have been analysed. The structures and chromosomal locations of 26 of the 30–50 elements estimated to be present in π2 have been determined. At one location two elements are inserted 100 base pairs (bp) apart, and in a second location two elements are only separated by the 8 bp duplicated upon P-element insertion. In addition to 2.9 kilobasepair (kbp) elements, elements with 14 different internal deletions from 1.3 to 2.3 kbp in size have been isolated. There are 7 copies of the 2–9 kbp element, 2 copies each of 5 internally deleted elements and a single copy of 9 internally deleted elements. One of the elements found twice is the KP element, which may play a role in the regulation of hybrid dysgenesis in strains which contain many copies of this element. Apart from internal deletions the elements are extremely homogeneous in DNA sequence, with only 2 single base polymorphisms detected twice each in over 16 kbp of P-element sequence. Although transpositions are infrequent in an inbred P cytotype strain such as π2, the distribution of these cloned elements indicates that when the genomic library was made, the strain was polymorphic with respect to element location. The distribution and structures of the element are discussed with respect to models for regulation of P-element transposition.

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