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Kirst Kingjones - One of the best experts on this subject based on the ideXlab platform.
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what goes up must come down transcription factors have their say in making Ecdysone pulses
Current Topics in Developmental Biology, 2013Co-Authors: Kirst KingjonesAbstract:Insect metamorphosis is one of the most fascinating biological processes in the animal kingdom. The dramatic transition from an immature juvenile to a reproductive adult is under the control of the steroid hormone Ecdysone, also known as the insect molting hormone. During Drosophila development, periodic pulses of Ecdysone are released from the prothoracic glands, upon which the hormone is rapidly converted in peripheral tissues to its biologically active form, 20-hydroxyEcdysone. Each hormone pulse has a unique profile and causes different developmental events, but we only have a rudimentary understanding of how the timing, amplitude, and duration of a given pulse are controlled. A key component involved in the timing of Ecdysone pulses is PTTH, a brain-derived neuropeptide. PTTH stimulates Ecdysone production through a Ras/Raf/ERK signaling cascade; however, comparatively little is known about the downstream targets of this pathway. In recent years, it has become apparent that transcriptional regulation plays a critical role in regulating the synthesis of Ecdysone, but only one transcription factor has a well-defined link to PTTH. Interestingly, many of the ecdysteroidogenic transcription factors were originally characterized as primary response genes in the Ecdysone signaling cascade that elicits the biological responses to the hormone in target tissues. To review these developments, we will first provide an overview of the transcription factors that act in the Drosophila Ecdysone regulatory hierarchy. We will then discuss the roles of these transcriptional regulators in controlling Ecdysone synthesis. In the last section, we will briefly outline transcription factors that likely have roles in regulating Ecdysone synthesis but have not been formally identified as downstream effectors of Ecdysone.
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nuclear receptor dhr4 controls the timing of steroid hormone pulses during drosophila development
PLOS Biology, 2011Co-Authors: Qiuxiang Ou, Adam Magico, Kirst KingjonesAbstract:In insects, precisely timed periodic pulses of the molting hormone Ecdysone control major developmental transitions such as molts and metamorphosis. The synthesis and release of Ecdysone, a steroid hormone, is itself controlled by PTTH (prothoracicotopic hormone). PTTH transcript levels oscillate with an 8 h rhythm, but its significance regarding the timing of Ecdysone pulses is unclear. PTTH acts on its target tissue, the prothoracic gland (PG), by activating the Ras/Raf/ERK pathway through its receptor Torso, however direct targets of this pathway have yet to be identified. Here, we demonstrate that Drosophila Hormone Receptor 4 (DHR4), a nuclear receptor, is a key target of the PTTH pathway and establishes temporal boundaries by terminating Ecdysone pulses. Specifically, we show that DHR4 oscillates between the nucleus and cytoplasm of PG cells, and that the protein is absent from PG nuclei at developmental times when low titer Ecdysone pulses occur. This oscillatory behavior is blocked when PTTH or torso function is abolished, resulting in nuclear accumulation of DHR4, while hyperactivating the PTTH pathway results in cytoplasmic retention of the protein. Increasing DHR4 levels in the PG can delay or arrest development. In contrast, reducing DHR4 function in the PG triggers accelerated development, which is caused by precocious Ecdysone signaling due to a failure to repress Ecdysone pulses. Finally, we show that DHR4 negatively regulates the expression of a hitherto uncharacterized cytochrome P450 gene, Cyp6t3. Disruption of Cyp6t3 function causes low ecdysteroid titers and results in heterochronic phenotypes and molting defects, indicating a novel role in the Ecdysone biosynthesis pathway. We propose a model whereby nuclear DHR4 controls the duration of Ecdysone pulses by negatively regulating Ecdysone biosynthesis through repression of Cyp6t3, and that this repressive function is temporarily overturned via the PTTH pathway by removing DHR4 from the nuclear compartment.
Yoshiaki Nakagawa - One of the best experts on this subject based on the ideXlab platform.
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arthropod nuclear receptors and their role in molting
FEBS Journal, 2009Co-Authors: Yoshiaki Nakagawa, Vincent C HenrichAbstract:The molting process in arthropods is regulated by steroid hormones acting via nuclear receptor proteins. The most common molting hormone is the ecdysteroid, 20-hydroxyEcdysone. The receptors of 20-hydroxyEcdysone have also been identified in many arthropod species, and the amino acid sequences determined. The functional molting hormone receptors consist of two members of the nuclear receptor superfamily, namely the Ecdysone receptor and the ultraspiracle, although the Ecdysone receptor may be functional, in some instances, without the ultraspiracle. Generally, the Ecdysone receptor/ultraspiracle heterodimer binds to a number of Ecdysone response elements, sequence motifs that reside in the promoter of various ecdysteroid-responsive genes. In the ensuing transcriptional induction, the Ecdysone receptor/ultraspiracle complex binds to 20-hydroxyEcdysone or to a cognate ligand that, in turn, leads to the release of a corepressor and the recruitment of coactivators. 3D structures of the ligand-binding domains of the Ecdysone receptor and the ultraspiracle have been solved for a few insect species. Ecdysone agonists bind to Ecdysone receptors specifically, and ligand–Ecdysone receptor binding is enhanced in the presence of the ultraspiracle in insects. The basic mode of ecdysteroid receptor action is highly conserved, but substantial functional differences exist among the receptors of individual species. Even though the transcriptional effects are apparently similar for ecdysteroids and nonsteroidal compounds such as diacylhydrazines, the binding shapes are different between them. The compounds having the strongest binding affinity to receptors ordinarily have strong molting hormone activity. The ability of the Ecdysone receptor/ ultraspiracle complex to manifest the effects of small lipophilic agonists has led to their use as gene switches for medical and agricultural applications.
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Molecular cloning of the Ecdysone receptor and the retinoid X receptor from the scorpion Liocheles australasiae.
FEBS Journal, 2007Co-Authors: Yoshiaki Nakagawa, Atsushi Sakai, Fumie Magata, Takehiko Ogura, Masahiro Miyashita, Hisashi MiyagawaAbstract:cDNAs of the Ecdysone receptor and the retinoid X receptor were cloned from the Japanese scorpion Liocheles australasiae, and the amino acid sequences were deduced. The full-length cDNA sequences of the L. australasiae Ecdysone receptor and the L. australasiae retinoid X receptor were 2881 and 1977 bp in length, respectively, and the open reading frames encoded proteins of 560 and 414 amino acids. The amino acid sequence of the L. australasiae Ecdysone receptor was similar to that of the Ecdysone receptor-A of the soft tick, Ornithodoros moubata (68%) and to that of the Ecdysone receptor-A1 of the lone star tick, Amblyomma americanum (66%), but showed lower similarity to the Ecdysone receptors of Orthoptera and Coleoptera (53–57%). The primary sequence of the ligand-binding region of the L. australasiae Ecdysone receptor was highly homologous to that of ticks (85–86%). The amino acid sequence of the L. australasiae retinoid X receptor was also homologous to the amino acid sequence of ultraspiracles of ticks (63%) and insects belonging to the orders Orthoptera and Coleoptera (60–64%). The identity of both the L. australasiae Ecdysone receptor and the L. australasiae retinoid X receptor to their lepidopteran and dipteran orthologs was less than 50%. The cDNAs of both the L. australasiae Ecdysone receptor (L. australasiae Ecdysone receptor-A) and the L. australasiae retinoid X receptor were successfully translated in vitro using a rabbit reticulocyte lysate system. An Ecdysone analog, ponasterone A, bound to L. australasiae Ecdysone receptor-A (KD = 4.2 nm), but not to L. australasiae retinoid X receptor. The L. australasiae retinoid X receptor did not enhance the binding of ponasterone A to L. australasiae Ecdysone receptor-A, although L. australasiae retinoid X receptor was necessary for the binding of L. australasiae Ecdysone receptor-A to Ecdysone response elements.
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Binding affinity of nonsteroidal Ecdysone agonists against the Ecdysone receptor complex determines the strength of their molting hormonal activity
European Journal of Biochemistry, 2003Co-Authors: Chieka Minakuchi, Yoshiaki Nakagawa, Manabu Kamimura, Hisashi MiyagawaAbstract:N-tert-Butyl-N,N'-dibenzoylhydrazine and its analogs are nonsteroidal Ecdysone agonists that exhibit insect molting hormonal and larvicidal activities. The interaction mode of those Ecdysone agonists with the heterodimer of the Ecdysone receptor and ultraspiracle has not been fully elucidated. We expressed the Ecdysone receptor B I and the ultraspiracle of the lepidopteran, Chilo suppressalis, using an in vitro transcription/translation system and confirmed, using gel-shift assays, that the proteins function as Ecdysone receptors. We also analyzed their ligand-binding affinity. A potent ecdysteroid, ponasterone A, specifically bound to the Ecdysone receptor with low affinity (K D = 55 nM), and the specific binding was dramatically increased (K D = 1.2 nM) in the presence of the ultraspiracle. For seven nonsteroidal Ecdysone agonists and five ecdysteroids, the binding activity to the in vitro-translated Ecdysone receptor-ultraspiracle complex was linearly correlated with the binding activity to the inherent receptor protein in the cell-free preparation of C. suppressalis integument. The binding to the Ecdysone receptor-ultraspiracle complex for a series of compounds was highly correlated with their molting hormonal activity, indicating that the binding affinity of nonsteroidal Ecdysone agonists to the Ecdysone receptor-ultraspiracle complex primarily determines the strength of their molting hormonal activity.
Peter Cherbas - One of the best experts on this subject based on the ideXlab platform.
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Ecdysone receptors and their biological actions.
Vitamins & Hormones, 2000Co-Authors: Lynn M. Riddiford, Peter Cherbas, James W. TrumanAbstract:Publisher Summary This chapter discusses the Ecdysone receptors and their biological actions. It also summarizes the insect endocrinology and the roles of these steroids in the molting and metamorphosis. Natural hormones that lead to molting and metamorphosis are Ecdysones. Molecules whose structures resemble those of the natural hormones are called ecdysteroids. The receptor for Ecdysone is a member of the nuclear receptor superfamily that acts as a ligand-dependent transcription factor. The vertebrate steroid hormone receptors act as homodimers whereas the functional Ecdysone receptor is always a heterodimer of receptor for Ecdysone (EcR) with another member of the nuclear receptor (NR) superfamily, Ultraspiracle, the insect homolog of the vertebrate retinoid X receptor (RXR). Two new technologies promise to transform the environment for investigations of insect hormones, Ecdysone receptor, and metamorphosis. The microarrays of expressed sequence tags are constructed (ESTs) and used hybridization to catalog changes in gene expression during metamorphosis. The first technological achievement is reviewed: the complete sequence of the Drosophila genome is obtained and is about to be released. It will be the first complete insect sequence and also the first genomic sequence from an organism that has served as a model for nuclear receptor endocrinology.
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Functional Ecdysone Receptor Is the Product of EcR and Ultraspiracle Genes
Nature, 1993Co-Authors: Tso-pang Yao, Lucy Cherbas, Peter Cherbas, Barry M. Forman, Ze-yu Jiang, Jasmine Chen, Michael Mckeown, Ronald M. EvansAbstract:Although the biological activity of the insect moulting hormone Ecdysone, is manifested through a hormonally regulated transcriptional cascade associated with chromosomal puffing, a direct association of the receptor with the puff has yet to be established. The cloned Ecdysone receptor (EcR) is by itself incapable of high-affinity DNA binding or transcriptional activation. Rather, these activities are dependent on heterodimer formation with Ultraspiracle (USP) the insect homologue of vertebrate retinoid X receptor. Here we report that native EcR and USP are co-localized on Ecdysone-responsive loci of polytene chromosomes. Moreover, we show that natural Ecdysones selectively promote physical association between EcR and USP, and conversely, that high-affinity hormone binding requires both EcR and USP. Replacement of USP with retinoid X receptor produces heterodimers with distinct pharmacological and functional properties. These results redefine the Ecdysone receptor as a dynamic complex whose activity may be altered by combinatorial interactions among subunits and ligand.
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The IVth Karlson Lecture: Ecdysone-responsive genes.
Insect biochemistry and molecular biology, 1993Co-Authors: Peter CherbasAbstract:Abstract Those of us who study Ecdysone action share at least two important long-range goals: (i) to understand the developmental specificity of steroid action in full molecular detail, by integrating Ecdysone action with our rapidly expanding knowledge of the molecular biology of insect development, and (ii) to better understand the nature of the steroid response and its evolution by taking advantage of the unparalleled opportunities for both genetic and comparative study afforded by the diversity of the “Ecdysone world”. However, until recently, the molecular fundamentals of the Ecdysone system were unknown and our efforts have, of necessity, been devoted to their elucidation. Now that the situation has changed: we have a small but varied catalog of Ecdysone-responsive genes for study and it is clear that some of these are tissue- and stage-specific in their expression. The Ecdysone receptor ( EcR ), like other steroid receptors a member of the nuclear receptor family, is now accessible to molecular study, and we have a preliminary understanding of the DNA sequences (EcREs) that bind receptor and specify a gene as Ecdysone-responsive. With these tools in hand and with the opportunity to turn to larger questions, it is a propitious moment to consider the nature of those questions and how Ecdysone can contribute to the answers.
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the drosophila ecr gene encodes an Ecdysone receptor a new member of the steroid receptor superfamily
Cell, 1991Co-Authors: Michael R Koelle, Peter Cherbas, Michael T. Bender, William S. Talbot, William A Segraves, David S. HognessAbstract:The steroid hormone Ecdysone triggers coordinate changes in Drosophila tissue development that result in metamorphosis. To advance our understanding of the genetic regulatory hierarchies controlling this tissue response, we have isolated and characterized a gene, EcR, for a new steroid receptor homolog and have shown that it encodes an Ecdysone receptor. First, EcR protein binds active ecdysteroids and is antigenically indistinguishable from the Ecdysone-binding protein previously observed in extracts of Drosophila cell lines and tissues. Second, EcR protein binds DNA with high specificity at Ecdysone response elements. Third, Ecdysone-responsive cultured cells express EcR, whereas Ecdysone-resistant cells derived from them are deficient in EcR. Expression of EcR in such resistant cells by transfection restores their ability to respond to the hormone. As expected, EcR is nuclear and found in all Ecdysone target tissues examined. Furthermore, the EcR gene is expressed at each developmental stage marked by a pulse of Ecdysone.
William A Segraves - One of the best experts on this subject based on the ideXlab platform.
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Ecdysone response genes govern egg chamber development during mid oogenesis in drosophila
Development, 1999Co-Authors: Michael Buszczak, Marc R Freeman, John R Carlson, Michael Bender, Lynn Cooley, William A SegravesAbstract:The steroid hormone Ecdysone regulates larval development and metamorphosis in Drosophila melanogaster through a complex genetic hierarchy that begins with a small set of early response genes. Here, we present data indicating that the Ecdysone response hierarchy also mediates egg chamber maturation during mid-oogenesis. E75, E74 and BR-C are expressed in a stage-specific manner while EcR expression is ubiquitous throughout oogenesis. Decreasing or increasing the ovarian Ecdysone titer using a temperature-sensitive mutation or exogenous Ecdysone results in corresponding changes in early gene expression. The stage 10 follicle cell expression of E75 in wild-type, K10 and EGF receptor (Egfr) mutant egg chambers reveals regulation of E75 by both the Egfr and Ecdysone signaling pathways. Genetic analysis indicates a germline requirement for Ecdysone-responsive gene expression. Germline clones of E75 mutations arrest and degenerate during mid-oogenesis and EcR germline clones exhibit a similar phenotype, demonstrating a functional requirement for Ecdysone responsiveness during the vitellogenic phase of oogenesis. Finally, the expression of Drosophila Adrenodoxin Reductase increases during mid-oogenesis and clonal analysis confirms that this steroidogenic enzyme is required in the germline for egg chamber development. Together these data suggest that the temporal expression profile of E75, E74 and BR-C may be a functional reflection of Ecdysone levels and that Ecdysone provides temporal signals regulating the progression of oogenesis and proper specification of dorsal follicle cell fates.
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Ecdysone response genes govern egg chamber development during mid oogenesis in drosophila
Development, 1999Co-Authors: Michael Buszczak, Marc R Freeman, John R Carlson, Michael Bender, Lynn Cooley, William A SegravesAbstract:The steroid hormone Ecdysone regulates larval development and metamorphosis in Drosophila melanogaster through a complex genetic hierarchy that begins with a small set of early response genes. Here, we present data indicating that the Ecdysone response hierarchy also mediates egg chamber maturation during mid-oogenesis. E75, E74 and BR-C are expressed in a stage-specific manner while EcR expression is ubiquitous throughout oogenesis. Decreasing or increasing the ovarian Ecdysone titer using a temperature-sensitive mutation or exogenous Ecdysone results in corresponding changes in early gene expression. The stage 10 follicle cell expression of E75 in wild-type, K10 and EGF receptor (Egfr) mutant egg chambers reveals regulation of E75 by both the Egfr and Ecdysone signaling pathways. Genetic analysis indicates a germline requirement for Ecdysone-responsive gene expression. Germline clones of E75 mutations arrest and degenerate during mid-oogenesis and EcR germline clones exhibit a similar phenotype, demonstrating a functional requirement for Ecdysone responsiveness during the vitellogenic phase of oogenesis. Finally, the expression of Drosophila Adrenodoxin Reductase increases during mid-oogenesis and clonal analysis confirms that this steroidogenic enzyme is required in the germline for egg chamber development. Together these data suggest that the temporal expression profile of E75, E74 and BR-C may be a functional reflection of Ecdysone levels and that Ecdysone provides temporal signals regulating the progression of oogenesis and proper specification of dorsal follicle cell fates.
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the drosophila ecr gene encodes an Ecdysone receptor a new member of the steroid receptor superfamily
Cell, 1991Co-Authors: Michael R Koelle, Peter Cherbas, Michael T. Bender, William S. Talbot, William A Segraves, David S. HognessAbstract:The steroid hormone Ecdysone triggers coordinate changes in Drosophila tissue development that result in metamorphosis. To advance our understanding of the genetic regulatory hierarchies controlling this tissue response, we have isolated and characterized a gene, EcR, for a new steroid receptor homolog and have shown that it encodes an Ecdysone receptor. First, EcR protein binds active ecdysteroids and is antigenically indistinguishable from the Ecdysone-binding protein previously observed in extracts of Drosophila cell lines and tissues. Second, EcR protein binds DNA with high specificity at Ecdysone response elements. Third, Ecdysone-responsive cultured cells express EcR, whereas Ecdysone-resistant cells derived from them are deficient in EcR. Expression of EcR in such resistant cells by transfection restores their ability to respond to the hormone. As expected, EcR is nuclear and found in all Ecdysone target tissues examined. Furthermore, the EcR gene is expressed at each developmental stage marked by a pulse of Ecdysone.
Lynn M. Riddiford - One of the best experts on this subject based on the ideXlab platform.
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Ecdysone receptors and their biological actions.
Vitamins & Hormones, 2000Co-Authors: Lynn M. Riddiford, Peter Cherbas, James W. TrumanAbstract:Publisher Summary This chapter discusses the Ecdysone receptors and their biological actions. It also summarizes the insect endocrinology and the roles of these steroids in the molting and metamorphosis. Natural hormones that lead to molting and metamorphosis are Ecdysones. Molecules whose structures resemble those of the natural hormones are called ecdysteroids. The receptor for Ecdysone is a member of the nuclear receptor superfamily that acts as a ligand-dependent transcription factor. The vertebrate steroid hormone receptors act as homodimers whereas the functional Ecdysone receptor is always a heterodimer of receptor for Ecdysone (EcR) with another member of the nuclear receptor (NR) superfamily, Ultraspiracle, the insect homolog of the vertebrate retinoid X receptor (RXR). Two new technologies promise to transform the environment for investigations of insect hormones, Ecdysone receptor, and metamorphosis. The microarrays of expressed sequence tags are constructed (ESTs) and used hybridization to catalog changes in gene expression during metamorphosis. The first technological achievement is reviewed: the complete sequence of the Drosophila genome is obtained and is about to be released. It will be the first complete insect sequence and also the first genomic sequence from an organism that has served as a model for nuclear receptor endocrinology.
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The Ecdysone receptor and ultraspiracle regulate the timing and progression of ovarian morphogenesis during Drosophila metamorphosis.
Development Genes and Evolution, 1998Co-Authors: Jason Hodin, Lynn M. RiddifordAbstract:Ecdysteroids regulate insect metamorphosis through the edysone receptor complex, a heterodimeric nuclear receptor consisting of the Ecdysone receptor (EcR) and its partner ultraspiracle (USP). Differentiation in the Drosophila ovary at metamorphosis correlates with colocalization of USP and the EcR-A isoform in all but one of eight mesoderm-derived somatic cell types. The one exception is the larval terminal filament (TF) cells, in which only USP is detectable during cell differentiation. In cells destined to form the basal stalks and anterior oviduct, USP colocalizes with what appears to be the EcR-B2 isoform. Flies heterozygous for a deletion of the EcR gene exhibit several defects in ovarian morphogenesis, including a heterochronic delay in the onset of terminal filament differentiation. Flies heterozygous for a strong usp allele exhibit accelerated TF differentiation. Flies simultaneously heterozygous for both EcR and usp have additional phenotypes, including several heterochronic shifts, delayed initiation and completion of terminal filament morphogenesis and delayed ovarian differentiation during the first day of metamorphosis. Terminal filament morphogenesis is severely disrupted in homozygous usp clones. Our results demonstrate that proper expression of the Ecdysone receptor complex is required to maintain the normal progression and timing of the events of ovarian differentiation in Drosophila. These findings are discussed in the context of a developmental and evolutionary role for the Ecdysone receptor complex in regulating the timing of ovarian differentiation in dipteran insects.