Ecdysone

Autocrine regulation of ecdysone synthesis by beta3-octopamine receptor in the prothoracic gland is essential for Drosophila metamorphosis.[Pubmed:25605909]

Proc Natl Acad Sci U S A. 2015 Feb 3;112(5):1452-7.

In Drosophila, pulsed production of the steroid hormone Ecdysone plays a pivotal role in developmental transitions such as metamorphosis. Ecdysone production is regulated in the prothoracic gland (PG) by prothoracicotropic hormone (PTTH) and insulin-like peptides (Ilps). Here, we show that monoaminergic autocrine regulation of Ecdysone biosynthesis in the PG is essential for metamorphosis. PG-specific knockdown of a monoamine G protein-coupled receptor, beta3-octopamine receptor (Octbeta3R), resulted in arrested metamorphosis due to lack of Ecdysone. Knockdown of tyramine biosynthesis genes expressed in the PG caused similar defects in Ecdysone production and metamorphosis. Moreover, PTTH and Ilps signaling were impaired by Octbeta3R knockdown in the PG, and activation of these signaling pathways rescued the defect in metamorphosis. Thus, monoaminergic autocrine signaling in the PG regulates Ecdysone biogenesis in a coordinated fashion on activation by PTTH and Ilps. We propose that monoaminergic autocrine signaling acts downstream of a body size checkpoint that allows metamorphosis to occur when nutrients are sufficiently abundant.

Ecdysone regulates morphogenesis and function of Malpighian tubules in Drosophila melanogaster through EcR-B2 isoform.[Pubmed:25476260]

Dev Biol. 2015 Feb 15;398(2):163-76.

Malpighian tubules are the osmoregulatory and detoxifying organs of Drosophila and its proper development is critical for the survival of the organism. They are made up of two major cell types, the ectodermal principal cells and mesodermal stellate cells. The principal and stellate cells are structurally and physiologically distinct from each other, but coordinate together for production of isotonic fluid. Proper integration of these cells during the course of development is an important pre-requisite for the proper functioning of the tubules. We have conclusively determined an essential role of Ecdysone hormone in the development and function of Malpighian tubules. Disruption of Ecdysone signaling interferes with the organization of principal and stellate cells resulting in malformed tubules and early larval lethality. Abnormalities include reduction in the number of cells and the clustering of cells rather than their arrangement in characteristic wild type pattern. Organization of F-actin and beta-tubulin also show aberrant distribution pattern. Malformed tubules show reduced uric acid deposition and altered expression of Na(+)/K(+)-ATPase pump. B2 isoform of Ecdysone receptor is critical for the development of Malpighian tubules and is expressed from early stages of its development.

Ecdysone signaling at metamorphosis triggers apoptosis of Drosophila abdominal muscles.[Pubmed:24051228]

Dev Biol. 2013 Nov 15;383(2):275-84.

One of the most dramatic examples of programmed cell death occurs during Drosophila metamorphosis, when most of the larval tissues are destroyed in a process termed histolysis. Much of our understanding of this process comes from analyses of salivary gland and midgut cell death. In contrast, relatively little is known about the degradation of the larval musculature. Here, we analyze the programmed destruction of the abdominal dorsal exterior oblique muscle (DEOM) which occurs during the first 24h of metamorphosis. We find that Ecdysone signaling through Ecdysone receptor isoform B1 is required cell autonomously for the muscle death. Furthermore, we show that the orphan nuclear receptor FTZ-F1, opposed by another nuclear receptor, HR39, plays a critical role in the timing of DEOM histolysis. Finally, we show that unlike the histolysis of salivary gland and midgut, abdominal muscle death occurs by apoptosis, and does not require autophagy. Thus, there is no set rule as to the role of autophagy and apoptosis during Drosophila histolysis.

INO80-dependent regression of ecdysone-induced transcriptional responses regulates developmental timing in Drosophila.[Pubmed:24468295]

Dev Biol. 2014 Mar 15;387(2):229-39.

Sequential pulses of the steroid hormone Ecdysone regulate the major developmental transitions in Drosophila, and the duration of each developmental stage is determined by the length of time between Ecdysone pulses. Ecdysone regulates biological responses by directly initiating target gene transcription. In turn, these transcriptional responses are known to be self-limiting, with mechanisms in place to ensure regression of hormone-dependent transcription. However, the biological significance of these transcriptional repression mechanisms remains unclear. Here we show that the chromatin remodeling protein INO80 facilitates transcriptional repression of Ecdysone-regulated genes during prepupal development. In ino80 mutant animals, inefficient repression of transcriptional responses to the late larval Ecdysone pulse delays the onset of the subsequent prepupal Ecdysone pulse, resulting in a significantly longer prepupal stage. Conversely, increased expression of ino80 is sufficient to shorten the prepupal stage by increasing the rate of transcriptional repression. Furthermore, we demonstrate that enhancing the rate of regression of the mid-prepupal competence factor betaFTZ-F1 is sufficient to determine the timing of head eversion and thus the duration of prepupal development. Although ino80 is conserved from yeast to humans, this study represents the first characterization of a bona fide ino80 mutation in any metazoan, raising the possibility that the functions of ino80 in transcriptional repression and developmental timing are evolutionarily conserved.

Ecdysone response elements in the distal promoter of the Bombyx Broad-Complex gene, BmBR-C.[Pubmed:24576019]

Insect Mol Biol. 2014 Jun;23(3):341-56.

The Bombyx mori silkworm's homologue of the Broad-Complex gene (BmBR-C) is transcribed from two promoters: a distal promoter (Pdist) and a proximal promoter (Pprox). As determined by a luciferase assay, the transcriptional activity of Pdist, but not Pprox, was activated by Ecdysone. Further analyses using reporters driven by sequential deletion Pdist mutants indicated that two regions, Ecdysone responsive element (EcRE)-D and EcRE-P, -4950 bp and -3480 bp upstream from the distal transcription start site, respectively, were important in the responsiveness of Pdist to 20-hydroxyEcdysone (20E); however, no significant sequence similarities were found between the canonical EcRE and the EcRE-D or EcRE-P regions. Electrophoretic mobility shift assays showed that both the EcRE-D and -P sequences specifically bound to Bombyx protein(s). Sequence analyses and competition assays suggested that the protein(s) bound to EcRE-P might include components other than the Ecdysone receptor (EcR), suggesting that BmBR-C transcription was indirectly activated by Ecdysone through the EcRE-P. Remarkably, protein binding to the mid-region of the EcRE-D, EcRE-Db, was competitively inhibited by an oligonucleotide containing the Drosophila hsp27 EcRE sequence. Furthermore, an anti-EcR antibody interfered with the formation of the protein-EcRE-Db complex. These results indicated that a functional Bombyx Ecdysone receptor binds to EcRE-D and activates the expression of BmBR-C.