Functional analysis of a naturally occurring variant dopa decarboxylase gene in Drosophila melanogaster using P element mediated germ line transformation

1987 ◽  
Vol 207 (2-3) ◽  
pp. 441-445 ◽  
Author(s):  
Zhi-Qing Chen ◽  
Ross B. Hodgetts
Keyword(s):  
Functional Analysis ◽  
Drosophila Melanogaster ◽  
Germ Line ◽  
P Element ◽  
Dopa Decarboxylase ◽  
Naturally Occurring ◽  
Germ Line Transformation

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.

The rough (ro+) gene as a dominant P-element marker in germ line transformation of Drosophila melanogaster

Gene ◽  
1992 ◽  
Vol 114 (2) ◽  
pp. 187-193 ◽  
Author(s):  
Trevor J. Lockett ◽  
Denise Lewy ◽  
Patricia Holmes ◽  
Kerrie Medveczky ◽  
Robert Saint
Keyword(s):  
Drosophila Melanogaster ◽  
Germ Line ◽  
P Element ◽  
Germ Line Transformation

scholarly journals cis-regulatory sequences responsible for alternative splicing of the Drosophila dopa decarboxylase gene.

1994 ◽  
Vol 14 (11) ◽  
pp. 7385-7393 ◽  
Author(s):  
J Shen ◽  
J Hirsh
Keyword(s):  
Alternative Splicing ◽  
Germ Line ◽  
Fusion Gene ◽  
P Element ◽  
Dopa Decarboxylase ◽  
Regulatory Sequences ◽  
Exon 1 ◽  
Alternatively Spliced ◽  
The Central Nervous System ◽  
Germ Line Transformation

The Drosophila dopa decarboxylase gene, Ddc, is expressed in the hypoderm and in specific sets of cells in the central nervous system (CNS). The unique Ddc primary transcript is alternatively spliced in these two tissues. The Ddc CNS mRNA contains all four exons (A through D), whereas the hypodermal mRNA contains only three exons (A, C, and D). To localize cis-regulatory sequences responsible for Ddc alternative splicing, a Ddc minigene and several fusion genes containing various amounts of Ddc sequences fused to fushi tarazu (ftz) exon 1 were constructed and introduced into flies by P-element-mediated germ line transformation. We find that Ddc intron ab and exon B are sufficient to regulate Ddc alternative splicing, since transcripts of a minimal fusion gene containing most of Ddc intron ab and exon B are spliced to exon B in the CNS but not in the hypoderm. These results indicate that Ddc alternative splicing is regulated by either a negative mechanism preventing splicing to exon B in the hypoderm or a positive mechanism activating splicing to exon B in the CNS. Our previous data suggest that Ddc hypodermal splicing is the actively regulated splicing pathway (J. Shen, C. J. Beall, and J. Hirsh, Mol. Cell. Biol. 13:4549-4555, 1993). Here we show that deletion of Ddc intron ab sequences selectively disrupts hypodermal splicing specificity. These results support a model in which Ddc alternative splicing is negatively regulated by a blockage mechanism preventing splicing to exon B in the hypoderm.

cis-regulatory sequences responsible for alternative splicing of the Drosophila dopa decarboxylase gene

1994 ◽  
Vol 14 (11) ◽  
pp. 7385-7393
Author(s):  
J Shen ◽  
J Hirsh
Keyword(s):  
Alternative Splicing ◽  
Germ Line ◽  
Fusion Gene ◽  
P Element ◽  
Dopa Decarboxylase ◽  
Regulatory Sequences ◽  
Exon 1 ◽  
Alternatively Spliced ◽  
The Central Nervous System ◽  
Germ Line Transformation

The Drosophila dopa decarboxylase gene, Ddc, is expressed in the hypoderm and in specific sets of cells in the central nervous system (CNS). The unique Ddc primary transcript is alternatively spliced in these two tissues. The Ddc CNS mRNA contains all four exons (A through D), whereas the hypodermal mRNA contains only three exons (A, C, and D). To localize cis-regulatory sequences responsible for Ddc alternative splicing, a Ddc minigene and several fusion genes containing various amounts of Ddc sequences fused to fushi tarazu (ftz) exon 1 were constructed and introduced into flies by P-element-mediated germ line transformation. We find that Ddc intron ab and exon B are sufficient to regulate Ddc alternative splicing, since transcripts of a minimal fusion gene containing most of Ddc intron ab and exon B are spliced to exon B in the CNS but not in the hypoderm. These results indicate that Ddc alternative splicing is regulated by either a negative mechanism preventing splicing to exon B in the hypoderm or a positive mechanism activating splicing to exon B in the CNS. Our previous data suggest that Ddc hypodermal splicing is the actively regulated splicing pathway (J. Shen, C. J. Beall, and J. Hirsh, Mol. Cell. Biol. 13:4549-4555, 1993). Here we show that deletion of Ddc intron ab sequences selectively disrupts hypodermal splicing specificity. These results support a model in which Ddc alternative splicing is negatively regulated by a blockage mechanism preventing splicing to exon B in the hypoderm.

P element mediated germ line transformation of Drosophila melanogaster with the Tc1 transposable DNA element from Caenorhabditis elegans

Genome ◽  
1994 ◽  
Vol 37 (3) ◽  
pp. 356-366 ◽  
Author(s):  
A. Alex Szekely ◽  
R. C. Woodruff ◽  
R. Mahendran
Keyword(s):  
Drosophila Melanogaster ◽  
Caenorhabditis Elegans ◽  
Germ Line ◽  
Molecular Data ◽  
P Element ◽  
P Elements ◽  
Line Transfer ◽  
Genomic Locations ◽  
Tc1 Element ◽  
Germ Line Transformation

Questions relating to the origin and regulation of mobile genetic elements are currently of considerable interest. Since it is now possible to address more precisely issues concerning the entry, dispersion, and regulation of elements within a virgin genome, one approach that may afford a better understanding of transposable elements in general could be provided by interspecific DNA transformation. Therefore, the Tc1 transposable DNA element from Caenorhabditis elegans was chosen as a proposed invading element of the Drosophila melanogaster genome. The basis for this selection resided in the inherent structural and functional similarities, as well as sequence identities, between the Caenorhabditis element and elements innate to Drosophila (e.g., P, HB1, and Uhu). Initial investigations were carried out to define a clone carrying an intact Tc1 element. This Tc1 element was inserted into a P transposon vector and two P–Tc1–ry+ constructs, differing only in insert orientation, were identified. P element mediated germ line transfer was then used to generate a transformant that was genetically and molecularly identified as containing a single, structurally intact Tc1 element at cytological location 64C4-5 on the third chromosome. The single P[(Tc1, ry+)]SAS-B insertion was thereafter mobilized by using a P[ry+Δ2-3] element as a transposase source, and the genetic and molecular data suggested that the insertion had been successfully reintegrated to a variety of genomic locations. On the basis of genetic and molecular analyses, the Tc1 element in the P[(Tc1, ry+)] transformed stock is not highly unstable in germ line and somatic tissues.Key words: Tc1 transposable element, transformation, Drosophila melanogaster, Caenorhabditis elegans, P elements.

scholarly journals 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 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.

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.

Transformation mapping of the regulatory elements of the ecdysone-inducible P1 gene of Drosophila melanogaster

1991 ◽  
Vol 11 (5) ◽  
pp. 2913-2917
Author(s):  
F Maschat ◽  
M L Dubertret ◽  
J A Lepesant
Keyword(s):  
Drosophila Melanogaster ◽  
Hormonal Regulation ◽  
Germ Line ◽  
Regulatory Elements ◽  
Fat Bodies ◽  
Germ Line Transformation ◽  
Third Instar Larvae

The transcription of the P1 gene is induced by 20-hydroxyecdysone in fat bodies of third-instar larvae. Germ line transformation showed that sequences between -138 to +276 contain elements required for a qualitatively correct developmental and hormonal regulation of P1 transcription. Sequences from -138 to -68 are essential for this expression.

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