The promoter region of the Drosophila alpha 2-tubulin gene directs testicular and neural specific expression

Development ◽  
1989 ◽  
Vol 106 (3) ◽  
pp. 581-587 ◽  
Author(s):  
J. Bo ◽  
P.C. Wensink
Keyword(s):  
Nervous System ◽  
Promoter Region ◽  
Germ Line ◽  
Fusion Gene ◽  
P Element ◽  
Tubulin Gene ◽  
Specific Expression ◽  
Spatial Expression Pattern ◽  
Males And Females ◽  
Temporal And Spatial

The temporal and spatial expression pattern of the Drosophila melanogaster alpha 2-tubulin gene (alpha 2) has been investigated by examining the expression of an alpha 2-lacZ fusion gene. When this fusion gene is introduced into the germ line by P-element mediated transformation, expression is only detected in chordotonal organs and testes. Chordotonal organs, which are sensory organs of the peripheral nervous system, express the gene from late embryonic through adult stages in both males and females. Testicular expression occurs from larval through adult stages and is limited to germ-line cells, the primary and secondary spermatocytes and perhaps the early spermatids.

scholarly journals P transposons controlled by the heat shock promoter.

1986 ◽  
Vol 6 (5) ◽  
pp. 1640-1649 ◽  
Author(s):  
H Steller ◽  
V Pirrotta
Keyword(s):  
Heat Shock ◽  
Germ Line ◽  
Fusion Gene ◽  
P Element ◽  
Translational Efficiency ◽  
Heat Shock Promoter ◽  
P Elements ◽  
Heat Shocks ◽  
Neomycin Resistance ◽  
Heat Shock Induction

We have transformed Drosophila melanogaster with modified P-element transposons, which express the transposase function from the heat-inducible hsp70 heat shock promoter. The Icarus transposon, which contains a direct hsp70-P fusion gene, behaved like a very active autonomous P element even before heat shock induction. Although heat shock led to abundant somatic transcription, transposition of the Icarus element was confined to germ line cells. To reduce the constitutive transposase activity observed for the Icarus element, we attenuated the translational efficiency of the transposase RNA by inserting the transposon 5 neomycin resistance gene between the hsp70 promoter and the P-element sequences. The resulting construct, called Icarus-neo, conferred resistance to G418, and its transposition was significantly stimulated by heat shock. Heat shocks applied during the embryonic or third instar larval stage had similar effects, indicating that transposition of P elements is not restricted to a certain developmental stage. Both Icarus and Icarus-neo destabilized snw in a P-cytotype background and thus at least partially overcome the repression of transposition. Our results suggest that the regulation of P-element transposition occurs at both the transcriptional and posttranscriptional levels.

scholarly journals Two independent cis-acting elements regulate the sex- and tissue-specific expression ofyp3inDrosophila melanogaster

1995 ◽  
Vol 66 (1) ◽  
pp. 9-17 ◽  
Author(s):  
Elaine Ronaldson ◽  
Mary Bownes
Keyword(s):  
Fat Body ◽  
Germ Line ◽  
Ovarian Follicle ◽  
Yolk Protein ◽  
P Element ◽  
Follicle Cells ◽  
Cell Type Specificity ◽  
Specific Expression ◽  
Cis Acting ◽  
Ideal System

SummaryInDrosophila, the threeyolk protein(yp) genes are transcribed in a sex-, tissue- and developmentally specific manner, providing an ideal system in which to investigate the factors involved in their regulation. The yolk proteins are synthesized in the fat body of adult females, and in the ovarian follicle cells surrounding the developing oocyte during stages 8–10 of oogenesis. We report here an analysis of theyolk protein 3(yp3) gene and its flanking sequences by means of P-element mediated germ-line transformation and demonstrate that a 747 bp promoter region is sufficient to direct sex-specific expression in the female fat body and both the temporal- and cell-type-specificity of expression during oogenesis. Two elements that independently governyp3transcription in these tissues have been separated and no other sequences in the upstream, downstream or coding regions have been identified that are autonomously involved inyp3expression.

scholarly journals A DNA segment controlling metal-regulated expression of the Drosophila melanogaster metallothionein gene Mtn

1987 ◽  
Vol 7 (5) ◽  
pp. 1710-1715
Author(s):  
E Otto ◽  
J M Allen ◽  
J E Young ◽  
R D Palmiter ◽  
G Maroni
Keyword(s):  
Drosophila Melanogaster ◽  
Mammalian Cells ◽  
Germ Line ◽  
Initiation Site ◽  
P Element ◽  
Specific Expression ◽  
Metallothionein Gene ◽  
Regulated Expression ◽  
Metallothionein Promoter ◽  
Simplex Virus

Cloned fragments of DNA including the Drosophila melanogaster metallothionein gene Mtn and different amounts of 5' flanking sequences were introduced into flies by P-element-mediated germ line transformation. Comparison of RNA levels in different transformants revealed that metal-regulated and tissue-specific expression of Mtn requires no more than 373 base pairs upstream of the initiation site of transcription. Transformants having an additional, transcribed copy of Mtn could tolerate increased concentrations of cadmium, indicating that Mtn expression is directly related to this phenotype. In separate experiments, these D. melanogaster promoter sequences were fused to the coding sequences of the herpes simplex virus thymidine kinase (TK) gene. After transfection of this fusion into baby hamster kidney cells, increases in TK activity and accumulation of TK RNA were inducible by metals. A series of 5' and 3' deletions showed that D. melanogaster sequences from -130 to -6 were sufficient to confer metal-regulated expression to the TK gene. The function of the D. melanogaster metallothionein promoter in mammalian cells indicates that the mechanism controlling metal regulation is evolutionarily conserved.

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 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.

scholarly journals The beta 3-tubulin gene of Drosophila melanogaster is essential for viability and fertility.

Genetics ◽  
1990 ◽  
Vol 126 (4) ◽  
pp. 991-1005
Author(s):  
M Kimble ◽  
R W Dettman ◽  
E C Raff
Keyword(s):  
Drosophila Melanogaster ◽  
Germ Line ◽  
Minor Component ◽  
Cell Types ◽  
P Element ◽  
Chromosome 2 ◽  
Tubulin Gene ◽  
Beta Tubulin ◽  
Cytoplasmic Microtubule ◽  
Tissue Organization

Abstract We have previously shown that the beta 3-tubulin gene of Drosophila melanogaster encodes a divergent isoform expressed in a complex developmental pattern. The beta 3 gene is transiently expressed in the embryo and again in the pupa at high levels in the developing musculature, and at lower levels in several different pupal tissues of ectodermal origin. Adult expression is confined to specific somatic cells in the gonads. In some of the cell types in which it is expressed, beta 3 is the sole or predominant beta-tubulin, while in others the beta 3 protein is a minor component of the beta-tubulin pool. The sites and timing of beta 3 expression demonstrated that beta 3-tubulin is utilized primarily in cytoplasmic microtubule arrays involved in changes in cell shape and tissue organization, and suggested to us that this isoform may be functionally specialized. To determine whether the expression of the beta 3 gene is essential for normal development, and to examine the specific functions of this divergent isoform, we have generated mutations within the gene. We determined that the small deficiency Df(2R)Px2, which deletes the 60C5,6-60D9,10 region of chromosome 2, removes all of the beta 3 coding sequences, and that the distal breakpoint of the deficiency is approximately 2 kb upstream from the start of transcription of the beta 3 gene. We have generated a total of 31 ethyl methanesulfonate- or diepoxybutane-induced recessive lethal or visible mutations which map within the deficiency. These mutations define 12 new lethal complementation groups, which together with two previously identified visible mutations, altogether identify 14 genes in this interval of the second chromosome. A lethal complementation group comprising mutations in the beta 3-tubulin gene (beta Tub60D) was identified by rescue of their lethality by a wild-type copy of the gene introduced into the genome via P element-mediated germ line transformation. Analysis of the homozygous and transheterozygous phenotypes of the five beta 3 mutations recovered (alleles designated B3t1-B3t5) demonstrates that beta 3-tubulin is essential for viability and fertility.

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.

The molecular manipulation of milk composition

Genome ◽  
1989 ◽  
Vol 31 (2) ◽  
pp. 950-955 ◽  
Author(s):  
A. J. Clark ◽  
S. Ali ◽  
A. L. Archibald ◽  
H. Bessos ◽  
P. Brown ◽  
...  
Keyword(s):  
Mammary Gland ◽  
Dna Sequences ◽  
Germ Line ◽  
Fusion Gene ◽  
Milk Proteins ◽  
Factor Ix ◽  
Farm Animals ◽  
Specific Expression ◽  
Human Factor Ix ◽  
Β Lactoglobulin

The introduction of cloned genes into the mouse germ line is now routine. Although more difficult technically, gene transfer has been accomplished in farm animals and offers the potential for genetic improvement. In this regard, we have been investigating the use of transgenic animals as production vehicles for high value proteins in milk. We have shown that DNA sequences derived from the gene encoding sheep β-lactoglobulin mediate efficient and specific expression in the mammary gland. A fusion gene comprising β-lactoglobulin sequences and those encoding antihemophilic human factor IX has been constructed. This construct has been introduced into sheep; it is expressed in the mammary gland, and the corresponding protein is secreted into milk.Key words: transgenic, mice, sheep, milk, proteins.

scholarly journals A DNA segment controlling metal-regulated expression of the Drosophila melanogaster metallothionein gene Mtn.

1987 ◽  
Vol 7 (5) ◽  
pp. 1710-1715 ◽  
Author(s):  
E Otto ◽  
J M Allen ◽  
J E Young ◽  
R D Palmiter ◽  
G Maroni
Keyword(s):  
Drosophila Melanogaster ◽  
Mammalian Cells ◽  
Germ Line ◽  
Initiation Site ◽  
P Element ◽  
Specific Expression ◽  
Metallothionein Gene ◽  
Regulated Expression ◽  
Metallothionein Promoter ◽  
Simplex Virus

Cloned fragments of DNA including the Drosophila melanogaster metallothionein gene Mtn and different amounts of 5' flanking sequences were introduced into flies by P-element-mediated germ line transformation. Comparison of RNA levels in different transformants revealed that metal-regulated and tissue-specific expression of Mtn requires no more than 373 base pairs upstream of the initiation site of transcription. Transformants having an additional, transcribed copy of Mtn could tolerate increased concentrations of cadmium, indicating that Mtn expression is directly related to this phenotype. In separate experiments, these D. melanogaster promoter sequences were fused to the coding sequences of the herpes simplex virus thymidine kinase (TK) gene. After transfection of this fusion into baby hamster kidney cells, increases in TK activity and accumulation of TK RNA were inducible by metals. A series of 5' and 3' deletions showed that D. melanogaster sequences from -130 to -6 were sufficient to confer metal-regulated expression to the TK gene. The function of the D. melanogaster metallothionein promoter in mammalian cells indicates that the mechanism controlling metal regulation is evolutionarily conserved.

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