Regulatory sequences of the Sgs-4 gene of Drosophila melanogaster analysed by P element-mediated transformation

Chromosoma ◽  
1987 ◽  
Vol 96 (1) ◽  
pp. 8-17 ◽  
Author(s):  
Annemarie Hofmann ◽  
Arne Keinhorst ◽  
Anton Krumm ◽  
G�nter Korge
Keyword(s):  
Drosophila Melanogaster ◽  
P Element ◽  
Regulatory Sequences

Delimiting regulatory sequences of the Drosophila melanogaster Ddc gene

1986 ◽  
Vol 6 (12) ◽  
pp. 4548-4557
Author(s):  
J Hirsh ◽  
B A Morgan ◽  
S B Scholnick
Keyword(s):  
Drosophila Melanogaster ◽  
Germ Line ◽  
Regulatory Elements ◽  
P Element ◽  
Developmental Expression ◽  
Upstream Region ◽  
Regulatory Sequences ◽  
Flanking Sequences

We delimited sequences necessary for in vivo expression of the Drosophila melanogaster dopa decarboxylase gene Ddc. The expression of in vitro-altered genes was assayed following germ line integration via P-element vectors. Sequences between -209 and -24 were necessary for normally regulated expression, although genes lacking these sequences could be expressed at 10 to 50% of wild-type levels at specific developmental times. These genes showed components of normal developmental expression, which suggests that they retain some regulatory elements. All Ddc genes lacking the normal immediate 5'-flanking sequences were grossly deficient in larval central nervous system expression. Thus, this upstream region must contain at least one element necessary for this expression. A mutated Ddc gene without a normal TATA boxlike sequence used the normal RNA start points, indicating that this sequences is not required for start point specificity.

scholarly journals Delimiting regulatory sequences of the Drosophila melanogaster Ddc gene.

1986 ◽  
Vol 6 (12) ◽  
pp. 4548-4557 ◽  
Author(s):  
J Hirsh ◽  
B A Morgan ◽  
S B Scholnick
Keyword(s):  
Drosophila Melanogaster ◽  
Germ Line ◽  
Regulatory Elements ◽  
P Element ◽  
Developmental Expression ◽  
Upstream Region ◽  
Regulatory Sequences ◽  
Flanking Sequences

We delimited sequences necessary for in vivo expression of the Drosophila melanogaster dopa decarboxylase gene Ddc. The expression of in vitro-altered genes was assayed following germ line integration via P-element vectors. Sequences between -209 and -24 were necessary for normally regulated expression, although genes lacking these sequences could be expressed at 10 to 50% of wild-type levels at specific developmental times. These genes showed components of normal developmental expression, which suggests that they retain some regulatory elements. All Ddc genes lacking the normal immediate 5'-flanking sequences were grossly deficient in larval central nervous system expression. Thus, this upstream region must contain at least one element necessary for this expression. A mutated Ddc gene without a normal TATA boxlike sequence used the normal RNA start points, indicating that this sequences is not required for start point specificity.

scholarly journals Definition of cis-acting elements regulating expression of the Drosophila melanogaster ninaE opsin gene by oligonucleotide-directed mutagenesis.

Genetics ◽  
1989 ◽  
Vol 121 (1) ◽  
pp. 77-87 ◽  
Author(s):  
D Mismer ◽  
G M Rubin
Keyword(s):  
Drosophila Melanogaster ◽  
The Body ◽  
Drosophila Virilis ◽  
P Element ◽  
Opsin Gene ◽  
Directed Mutagenesis ◽  
Homologous Gene ◽  
Regulatory Sequences ◽  
Specific Expression ◽  
Cis Acting

Abstract We have analyzed the cis-acting regulatory sequences of the Rh1 (ninaE) gene in Drosophila melanogaster by P-element-mediated germline transformation of indicator genes transcribed from mutant ninaE promoter sequences. We have previously shown that a 200-bp region extending from -120 to +67 relative to the transcription start site is sufficient to obtain eye-specific expression from the ninaE promoter. In the present study, 22 different 4-13-bp sequences in the -120/+67 promoter region were altered by oligonucleotide-directed mutagenesis. Several of these sequences were found to be required for proper promoter function; two of these are conserved in the promoter of the homologous gene isolated from the related species Drosophila virilis. Alteration of a conserved 9-bp sequence results in aberrant, low level expression in the body. Alteration of a separate 11-bp sequence, found in the promoter regions of several photoreceptor-specific genes of Drosophila, results in an approximately 15-fold reduction in promoter efficiency but without apparent alteration of tissue-specificity. A protein factor capable of interacting with this 11-bp sequence has been detected by DNaseI footprinting in embryonic nuclear extracts. Finally, we have further characterized two separable enhancer sequences previously shown to be required for normal levels of expression from this promoter.

Analysis of the promoter of the Rh2 opsin gene in Drosophila melanogaster.

Genetics ◽  
1988 ◽  
Vol 120 (1) ◽  
pp. 173-180
Author(s):  
D Mismer ◽  
W M Michael ◽  
T R Laverty ◽  
G M Rubin
Keyword(s):  
Drosophila Melanogaster ◽  
Fusion Gene ◽  
Photoreceptor Cells ◽  
Histochemical Staining ◽  
P Element ◽  
Opsin Gene ◽  
Regulatory Sequences ◽  
Specific Expression ◽  
Beta Galactosidase ◽  
Cat Expression

Abstract We have analyzed the cis-acting regulatory sequences of the Drosophila melanogaster Rh2 gene that encodes the protein component of a rhodopsin which is expressed in ocellar photoreceptor cells. DNA fragments containing the start point of transcription of the Rh2 gene were fused to either the Escherichia coli chloramphenicol acetyltransferase (CAT) or lacZ (beta-galactosidase) genes and introduced into the Drosophila germline by P-element-mediated transformation. Expression of the E. coli genes was then used to assay the ability of various sequences from the Rh2 gene to confer upon the indicator genes the Rh2 pattern of expression. Fragments containing between 4.3 kb and 183 bp upstream of the start of transcription plus the first 32 bp of the 5'-untranslated leader were found to result in nearly identical levels of head-specific CAT expression. Deletion of Rh2 sequences distal to position -112 bp resulted in loss of detectable CAT expression from these Rh2/CAT fusion constructs. We have, therefore, defined a region essential for head-specific expression of the Rh2 gene to a region extending from -183 to -112. We have determined the DNA sequence of the Rh2 promoter from -448 to +32 and have found an 11-bp sequence which is also present in the upstream flanking sequences of two other photoreceptor-specific genes (ninaE and ninaC). By histochemical staining of beta-galactosidase expressed under the control of the Rh2 promoter and by analyzing the effect of the ocelliless mutation on the expression of an Rh2/CAT fusion gene, we have been able to demonstrate that this promoter is active in ocelli.

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.

scholarly journals A hobo Transgene That Encodes the P-Element Transposase in Drosophila melanogaster: Autoregulation and Cytotype Control of Transposase Activity

Genetics ◽  
2002 ◽  
Vol 161 (1) ◽  
pp. 195-204 ◽  
Author(s):  
Michael J Simmons ◽  
Kevin J Haley ◽  
Craig D Grimes ◽  
John D Raymond ◽  
Jarad B Niemi
Keyword(s):  
Drosophila Melanogaster ◽  
Gonadal Dysgenesis ◽  
P Element ◽  
Hsp70 Gene ◽  
Alternate Splicing ◽  
Male And Female ◽  
P Elements ◽  
Male Germline ◽  
Female Germline ◽  
Transposase Expression

Abstract Drosophila were genetically transformed with a hobo transgene that contains a terminally truncated but otherwise complete P element fused to the promoter from the Drosophila hsp70 gene. Insertions of this H(hsp/CP) transgene on either of the major autosomes produced the P transposase in both the male and female germlines, but not in the soma. Heat-shock treatments significantly increased transposase activity in the female germline; in the male germline, these treatments had little effect. The transposase activity of two insertions of the H(hsp/CP) transgene was not significantly greater than their separate activities, and one insertion of this transgene reduced the transposase activity of P(ry+, Δ2-3)99B, a stable P transgene, in the germline as well as in the soma. These observations suggest that, through alternate splicing, the H(hsp/CP) transgene produces a repressor that feeds back negatively to regulate transposase expression or function in both the somatic and germline tissues. The H(hsp/CP) transgenes are able to induce gonadal dysgenesis when the transposase they encode has P-element targets to attack. However, this ability and the ability to induce P-element excisions are repressed by the P cytotype, a chromosomal/cytoplasmic state that regulates P elements in the germline.

scholarly journals The Regulatory Properties of Autonomous Subtelomeric P Elements Are Sensitive to a Suppressor of variegation in Drosophila melanogaster

Genetics ◽  
1996 ◽  
Vol 143 (4) ◽  
pp. 1663-1674 ◽  
Author(s):  
Stéphane Ronsseray ◽  
Monique Lehmann ◽  
Danielle Nouaud ◽  
Dominique Anxolabéhère
Keyword(s):  
Drosophila Melanogaster ◽  
Genetic Recombination ◽  
Natural Populations ◽  
P Element ◽  
Single Element ◽  
Heterochromatin Protein 1 ◽  
X Chromosomes ◽  
P Elements ◽  
Associated Sequences ◽  
Regulatory Properties

Abstract Genetic recombination was used in Drosophila melanogaster to isolate P elements, inserted at the telomeres of X chromosomes (cytological site 1A) from natural populations, in a genetic background devoid of other P elements. We show that complete maternally inherited P repression in the germline (P cytotype) can be elicited by only two autonomous P elements at 1A and that a single element at this site has partial regulatory properties. The analysis of the surrounding chromosomal regions of the P elements at 1A shows that in all cases these elements are flanked by Telomeric Associated Sequences, tandemly repetitive noncoding sequences that have properties of heterochromatin. In addition, we show that the regulatory properties of P elements at 1A can be inhibited by some of the mutant alleles of the Su(var)205 gene and by a deficiency of this gene. However, the regulatory properties of reference P strains (Harwich and Texas 007) are not impaired by Su(var)205 mutations. Su(var)205 encodes Heterochromatin Protein 1 (HP1). These results suggest that the HP1 dosage effect on the P element properties is sitedependent and could involve the structure of the chromatin.

scholarly journals hobo enhancer trapping mutagenesis in Drosophila reveals an insertion specificity different from P elements.

Genetics ◽  
1993 ◽  
Vol 135 (4) ◽  
pp. 1063-1076 ◽  
Author(s):  
D Smith ◽  
J Wohlgemuth ◽  
B R Calvi ◽  
I Franklin ◽  
W M Gelbart
Keyword(s):  
Drosophila Melanogaster ◽  
Transposable Element ◽  
Enhancer Trap ◽  
P Element ◽  
Present Evidence ◽  
P Elements ◽  
Element Insertion ◽  
Enhancer Trapping ◽  
Indispensable Tool

Abstract P element enhancer trapping has become an indispensable tool in the analysis of the Drosophila melanogaster genome. However, there is great variation in the mutability of loci by these elements such that some loci are relatively refractory to insertion. We have developed the hobo transposable element for use in enhancer trapping and we describe the results of a hobo enhancer trap screen. In addition, we present evidence that a hobo enhancer trap element has a pattern of insertion into the genome that is different from the distribution of P elements in the available database. Hence, hobo insertion may facilitate access to genes resistant to P element insertion.

scholarly journals Genetic and Molecular Characterization of sting, a Gene Involved in Crystal Formation and Meiotic Drive in the Male Germ Line of Drosophila melanogaster

Genetics ◽  
1999 ◽  
Vol 151 (2) ◽  
pp. 749-760 ◽  
Author(s):  
Armin Schmidt ◽  
Gioacchino Palumbo ◽  
Maria P Bozzetti ◽  
Patrizia Tritto ◽  
Sergio Pimpinelli ◽  
...  
Keyword(s):  
Amino Acids ◽  
Drosophila Melanogaster ◽  
Null Allele ◽  
Germ Line ◽  
Meiotic Drive ◽  
P Element ◽  
Crystal Formation ◽  
Male Sterile ◽  
Male Sterile Mutation

Abstract The sting mutation, caused by a P element inserted into polytene region 32D, was isolated by a screen for male sterile insertions in Drosophila melanogaster. This sterility is correlated with the presence of crystals in spermatocytes and spermatids that are structurally indistinguishable from those produced in males carrying a deficiency of the Y-linked crystal (cry) locus. In addition, their morphology is needle-like in Ste+ flies and star-shaped in Ste flies, once again as observed in cry– males. The sti mutation leads to meiotic drive of the sex chromosomes, and the strength of the phenomenon is correlated with the copy number of the repetitive Ste locus. The same correlation is also true for the penetrance of the male sterile mutation. A presumptive sti null allele results in male sterility and lethal maternal effect. The gene was cloned and shown to code for a putative protein that is 866 amino acids long. A C-terminal domain of 82 amino acids is identified that is well conserved in proteins from different organisms. The gene is expressed only in the germline of both sexes. The interaction of sting with the Ste locus can also be demonstrated at the molecular level. While an unprocessed 8-kb Ste primary transcript is expressed in wild-type males, in X/Y homozygous sti males, as in X/Y cry– males, a 0.7-kb mRNA is produced.

Identification of Chromosome Inheritance Modifiers in Drosophila melanogaster

Genetics ◽  
2001 ◽  
Vol 157 (4) ◽  
pp. 1623-1637 ◽  
Author(s):  
Kenneth W Dobie ◽  
Cameron D Kennedy ◽  
Vivienne M Velasco ◽  
Tory L McGrath ◽  
Juliani Weko ◽  
...  
Keyword(s):  
Drosophila Melanogaster ◽  
Biological Activity ◽  
Cancer Progression ◽  
Birth Defects ◽  
Mitotic Chromosome ◽  
P Element ◽  
Inverse Pcr ◽  
Gtpase Activating Protein ◽  
New Genes ◽  
Genomic Analyses

Abstract Faithful chromosome inheritance is a fundamental biological activity and errors contribute to birth defects and cancer progression. We have performed a P-element screen in Drosophila melanogaster with the aim of identifying novel candidate genes involved in inheritance. We used a “sensitized” minichromosome substrate (J21A) to screen ∼3,000 new P-element lines for dominant effects on chromosome inheritance and recovered 78 Sensitized chromosome inheritance modifiers (Scim). Of these, 69 decreased minichromosome inheritance while 9 increased minichromosome inheritance. Fourteen mutations are lethal or semilethal when homozygous and all exhibit dramatic mitotic defects. Inverse PCR combined with genomic analyses identified P insertions within or close to genes with previously described inheritance functions, including wings apart-like (wapl), centrosomin (cnn), and pavarotti (pav). Further, lethal insertions in replication factor complex 4 (rfc4) and GTPase-activating protein 1 (Gap1) exhibit specific mitotic chromosome defects, discovering previously unknown roles for these proteins in chromosome inheritance. The majority of the lines represent mutations in previously uncharacterized loci, many of which have human homologs, and we anticipate that this collection will provide a rich source of mutations in new genes required for chromosome inheritance in metazoans.

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