The role of the BR-C locus on the expression of genes located at the ecdysone-regulated 3C puff of Drosophila melanogaster (original) (raw)
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Genetical Research, 1992
SummaryThe Drosophila melanogasterecd1 mutation causes a severe temperature-sensitive deficiency in the titre of the steroid hormone eedysone. This mutation was used to investigate the role of eedysone in both the transcription of the genes mapped at the 3C11–12 intermoult puff region and the puff formation. Thoroughly synchronized ecd1 larvae were shifted to the non-permissive temperature at various times of the development; after 24 or 48 h, the levels of the transcripts derived from Sgs-4, Pig-1 and ng-1, the three genes located at the 3C11–12 polytene bands, were determined. The results showed that the levels of the transcripts encoded by Pig-1 and ng-1 are unaffected by the drop in the ecdysone titre occurring in non-permissive conditions whereas the amount of Sgs-4 mRNA is greatly reduced. These data clearly indicate that transcription of the three genes mapped within the puff region is affected differently by the hormone. Furthermore, ecd1 larvae cultured at the non-permissiv...
Development, 1993
During Drosophila third instar larval development, one or more pulses of the steroid hormone ecdysone activate three temporally distinct sets of genes in the salivary glands, represented by puffs in the polytene chromosomes. The intermolt genes are induced first, in mid-third instar larvae; these genes encode a protein glue used by the animal to adhere itself to a solid substrate for metamorphosis. The intermolt genes are repressed at puparium formation as a high titer ecdysone pulse directly induces a small set of early regulatory genes. The early genes both repress their own expression and activate more than 100 late secondary-response genes. The Broad-Complex (BR-C) is an early ecdysone-inducible gene that encodes a family of DNA binding proteins defined by at least three lethal complementation groups: br, rbp, and l(1)2Bc. We have found that the BR-C is critical for the appropriate regulation of all three classes of ecdysone-inducible genes. Both rbp and l(1)2Bc are required for...
The EMBO journal, 1994
In Drosophila, all of the major metamorphic transitions are regulated by changes in the titer of the steroid hormone ecdysone. Here we examine how a key regulator of metamorphosis and primary ecdysone response gene, the Broad-Complex, transmits the hormonal signal to one of its targets, the Sgs-4 glue gene. We show that Broad-Complex RNAs accumulate in mid third instar larval salivary glands prior to Sgs-4 induction, as expected for the products of a gene that regulates the timing of Sgs-4 activation. The Broad-Complex codes for a family of zinc finger transcriptional regulators. We have identified a number of binding sites for these proteins in sequences known to regulate the timing of Sgs-4 induction, and have used these sites to derive a binding consensus for each protein. Some of these binding sites are required in vivo for Sgs-4 activity. In addition, rbp+, a genetically defined Broad-Complex function that is required for Sgs-4 induction, acts through these Broad-Complex bindin...
Developmental Biology, 2000
Metamorphosis in Drosophila melanogaster is orchestrated by the steroid hormone ecdysone, which triggers a cascade of primary-response transcriptional regulators and secondary effector genes during the third larval instar and prepupal periods of development. The early ecdysone-response Broad-Complex (BR-C) gene, a key regulator of this cascade, is defined by three complementing functions (rbp, br, and 2Bc) and encodes several distinct zinc-finger-containing isoforms (Z1 to Z4). Using isoform-specific polyclonal antibodies we observe in the fat body a switch in BR-C isoform expression from the Z2 to the other three isoforms during the third instar. We show that the 2Bc ؉ function that corresponds presumably to the Z3 isoform is required for the larval fat body-specific expression of a transgenic construct (AE) in which the lacZ gene is under the control of the ecdysone-regulated enhancer and minimal promoter of the fat body protein 1 (Fbp1) gene. Using hs(BR-C) transgenes, we demonstrate that overexpression of Z1, Z3, or Z4, but not Z2, is able to rescue AE activity with faithful tissue specificity in a BR-C null (npr1) genetic context, demonstrating a partial functional redundancy between Z1, Z3, and Z4 isoforms. We also show that continuous overexpression of Z2 during the third instar represses AE, while conversely, expression of Z3 earlier than its normal onset induces precocious expression of the construct. This finding establishes a tight correlation between the dynamic pattern of expression of the BR-C isoforms and their individual repressive or inductive roles in AE regulation. Altogether our results demonstrate that the balance between BR-C protein isoforms in the fat body mediates, in part, the precise timing of the ecdysone activation of the AE construct but does not modulate its tissue specificity.
Journal of Molecular Biology, 1986
We have determined the molecular organization of an ecdysterone-responsive puff site in Drosophila melanogaster. The 71E puff site contains a tightly linked cluster of at least seven genes within a neighborhood of 10 x 103 base-pairs. All the genes are expressed in a tissuespecific manner in either the larval or the prepupal salivary gland. However, these genes can be divided into two groups on the basis of their temporal pattern of transcription. Six of the genes are expressed only in prepupal salivary glands and are arranged as three divergently transcribed pairs. Nestled within this region is one gene expressed primarily in late third-instar salivary glands. We conclude that this developmentally complex puff site contains six members of the ecdysterone-induced "late"-gene set and one member of the ecdysterone-regulated "intermolt"-gene set. Additional complexity is found at the transcript level: a heterogeneously sized population of RNA molecules arises fi'om each of the seven genes.
Journal of Molecular Biology, 1986
We have determined the molecular organization of an ecdysterone-responsive puff site in Drosophila melanogaster. The 71E puff site contains a tightly linked cluster of at least seven genes within a neighborhood of 10 x 103 base-pairs. All the genes are expressed in a tissuespecific manner in either the larval or the prepupal salivary gland. However, these genes can be divided into two groups on the basis of their temporal pattern of transcription. Six of the genes are expressed only in prepupal salivary glands and are arranged as three divergently transcribed pairs. Nestled within this region is one gene expressed primarily in late third-instar salivary glands. We conclude that this developmentally complex puff site contains six members of the ecdysterone-induced "late"-gene set and one member of the ecdysterone-regulated "intermolt"-gene set. Additional complexity is found at the transcript level: a heterogeneously sized population of RNA molecules arises fi'om each of the seven genes.
Specific transcriptional responses to juvenile hormone and ecdysone in Drosophila
Insect Biochemistry and Molecular Biology, 2007
Previous studies have shown that ecdysone (E), and its immediate downstream product 20-hydroxyecdysone (20E), can have different biological functions in insects, suggesting that E acts as a distinct hormone. Here, we use Drosophila larval organ culture in combination with microarray technology to identify genes that are transcriptionally regulated by E, but which show little or no response to 20E. These genes are coordinately expressed for a brief temporal interval at the onset of metamorphosis, suggesting that E acts together with 20E to direct puparium formation. We also show that E74B, pepck, and CG14949 can be induced by juvenile hormone III (JH III) in organ culture, and that CG14949 can be induced by JH independently of protein synthesis. In contrast, E74A and E75A show no response to JH in this system. These studies demonstrate that larval organ culture can be used to identify Drosophila genes that are regulated by hormones other than 20E, and provide a basis for studying crosstalk between multiple hormone signaling pathways. r
Genome, 2001
The effect of ecdysone on the puffing activity of the polytene chromosomes of Ceratitis capitata has been studied in organ cultures of late-larval salivary glands. Culture of glands from 120-h-old larvae (puff stage 1) in the presence of ecdysone resulted in the initiation of the late-larval puffing cycle that is normally observed in 145-h-old larvae (puff stage 4). During a 7-h period in the presence of ecdysone, the puffing patterns of most loci resembled the in vivo patterns observed in the period between puff stages 4 and 10, indicating that the first puffing cycle can be initiated by the hormone and proceed almost to completion, in vitro. Culture of salivary glands in the presence of ecdysone and a protein-synthesis inhibitor, as well as ecdysone withdrawal and readdition experiments, indicated that most of the ecdysone-regulated puffs could be categorized into three classes: (i) the puffs that were suppressed immediately by ecdysone, even in the absence of protein synthesis; (ii) the puffs that were induced directly by ecdysone; and (iii) the puffs that were induced indirectly by ecdysone, that is, they were induced after a lag period of a few hours and required protein synthesis for their induction.
Journal of Molecular Biology, 1996
Early metamorphic development in Drosophila melanogaster is initiated by pulses of the steroid hormone ecdysone, which are transduced into University of Pennsylvania tissue-specific transcriptional cascades. This process begins with the Philadelphia, PA 19104-6018 hormone-dependent activation of a set of transcription factors (early genes) USA that, in turn, activate set of tissue-specific effector genes (late genes). The 2 Howard Hughes Medical 71E cytogenetic region of the salivary gland polytene genome contains Institute, Department of several ecdysone-regulated transcription units. Molecular techniques were Human Genetics, 5200 Eccles used to analyze these genes, their transcriptional program and their Institute of Human Genetics evolutionary relatedness. We find that this region contains a cluster of ten University of Utah, Salt Lake coordinately regulated late genes (L71 genes) that are organized as five City, UT 84112, USA divergently transcribed gene pairs. Maximum parsimony analysis suggests that an ancestral L71 gene duplicated to form the first gene pair which was, in turn, duplicated to form the set of gene pairs. The L71 gene products form a family of small, chemically basic proteins with a conserved backbone of cysteine residues. In addition, the 71E region contains another gene (I71-1) with the regulatory and biochemical characteristics of the salivary gland intermolt glue proteins.
Developmental Biology, 2000
The pulse of ecdysone that triggers Drosophila metamorphosis activates six early genes in a primary response made visible by polytene chromosome puffs. The secondary response is detected by the induction of over 100 late puffs, only a few of which have been subject to molecular genetic analysis. We present a molecular and mutational analysis of the L63 gene responsible for the late puff at 63E. This gene contains overlapping L63A, B, and C transcription units of which the A unit encodes two isoforms and the B unit three. The C unit, which exhibits little activity, encodes one of the B isoforms. Evidence that L63B, but not L63A, transcription is ecdysone responsive derives from their developmental transcription profiles and from P-element mutagenesis showing that ecdysone induction of the 63E puff requires sequences adjacent to the 5 end of L63B but not those adjacent to the 5 end of L63A. L63-specific lethal mutations showed that L63 is required not only for metamorphosis, but also maternally and for embryonic and larval development. The L63 proteins contain a common C-terminal 294-aa sequence that is 71% identical to the CDK sequence of the murine PFTAIRE protein. In vivo tests of L63 proteins altered by site-directed mutagenesis showed that they exhibit CDK functions. L63 proteins are widely distributed among late larval and prepupal tissues and are unlikely to be involved in cell cycle functions.