The uptake and release of ponasterone a by the Kc cell line of Drosophila melanogaster

Beckers, C.; Maróy, Péter; Dennis, Roger D.; O’Connor, John D.; Emmerich, Hans

doi:10.1016/0303-7207(80)90103-3

Cited by 34 publications

(10 citation statements)

References 17 publications

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“…In contrast, we used a constant radiolabeled ligand concentration and a variable competitor concentration. The competition binding assay confirmed the affinity of 20E and tebufenozide for the EcRs as demonstrated by many other investigators who proved that both bisacylhydrazines and 20E competitively displaced 3 H-PoA (Beckers et al, 1980;Wing, 1988;Spindler-Barth et al 1991;Quack et al, 1995;Sobek et al, 1993;Mikitani, March 2005 1996; Hsu et al, 1997). Competitive displacement of 3 H-PoA was also used in intact imaginal wing discs from lepidopteran and coleopteran larvae (Smagghe et al, 1996(Smagghe et al, , 1999.…”

Section: Discussionsupporting

confidence: 80%

See 1 more Smart Citation

Action of 24-epibrassinolide on a cell line of the beet armyworm,Spodoptera exigua

Decombel

Tirry

Smagghe

2005

Arch. Insect Biochem. Physiol.

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The Spodoptera exigua cell line Se4 is sensitive for ecdysteroid activity stimulated by the insect molting hormone, 20-hydroxyecdysone (20E), showing a cease in cell proliferation (with 50% inhibition around 1 microM) and characteristic cell morphology changes with aggregation and formation of long filamentous cytoplasmic extensions. The bisacylhydrazine tebufenozide also triggered such typical cellular effects in Se4, and in addition, it showed an affinity for binding in competition with 3H-ponasterone A (PoA) that was similar to 20E (with 50% competition around 1 microM), confirming that such non-ecdysteroids display an ecdysteroid agonist activity. In contrast, when Se4 cells were incubated with the native plant hormone 24-epibrassinolide (24BR), none of the effects triggered by 20E were observed. Hence, a competition binding experiment with 3H-PoA demonstrated no affinity of 24BR for binding to the ecdysteroid receptor in the Se4 cell line. In another series of experiments, the Se4 cell line was tested in sensitivity response to increased acetylcholinesterase (AchE) activity after treatment with ecdysteroid active compounds. The AchE activity measured in the cell line is discussed in relation to inhibition by eserine. The obtained results suggest that 24BR exerted no ecdysteroid activity.

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Section: Discussionsupporting

confidence: 80%

“…The competition binding experiments were performed according to Beckers et al (1980) and made use of 3 H-PoA (8 nM) incubated with various concentrations of cold competitor. For this experiment, cells were harvested 6 days after subculturing and centrifuged at room temperature at 600g for 5 min.…”

Section: Competition Binding Experimentsmentioning

confidence: 99%

Action of 24-epibrassinolide on a cell line of the beet armyworm,Spodoptera exigua

Decombel

Tirry

Smagghe

2005

Arch. Insect Biochem. Physiol.

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“…The key steroid hormone in arthropods is ecdysterone (20-hydroxyecdysone), which regulates molt cycles and is also involved in other processes, such as reproduction, etc. Nuclear receptors for ecdysterone in arthropods, including Drosophila melanogaster, have been identified previously (13,17,63,82,83,102). It will be important to study these receptors at the molecular level to compare mechanisms of hormonal gene regulation in widely divergent organisms as well as to obtain a better understanding of mechanisms underlying major developmental events in insects, such as pupation, etc.…”

mentioning

confidence: 99%

Ecdysterone receptor is a sequence-specific transcription factor involved in the developmental regulation of heat shock genes.

Luo¹,

Amin²,

Voellmy³

1991

Mol. Cell. Biol.

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Purification of ecdysterone receptor from Drosophila melanogaster to apparent homogeneity is reported. Purified receptor binds specifically to several sequences in the promoters of the developmentally active hsp27 and hsp23 heat shock genes that were previously implied in ecdysterone regulation of the genes and that share limited homology among themselves and with mammalian steroid receptor binding sites. Some of these elements confer ecdysterone regulation on a basal promoter in transfected cells, acting in a synergistic fashion. Transcription in vitro of promoters containing such elements is stimulated up to 100-fold by added purified ecdysterone receptor, depending on receptor dosage and the number of elements present. Transcriptional enhancement requires sequence-specific binding of receptor to template promoters which facilitates the formation of a preinitiation complex. Ecdysterone stimulates DNA binding of the receptor in vitro.A wealth of information concerning steroid hormone receptors and mechanisms of steroid hormone regulation in vertebrate systems has accumulated (10,11,29,34,39,72,101). In contrast, very little is known about steroid hormone receptors in lower eukaryotes. The key steroid hormone in arthropods is ecdysterone (20-hydroxyecdysone), which regulates molt cycles and is also involved in other processes, such as reproduction, etc. Nuclear receptors for ecdysterone in arthropods, including Drosophila melanogaster, have been identified previously (13,17,63,82,83,102). It will be important to study these receptors at the molecular level to compare mechanisms of hormonal gene regulation in widely divergent organisms as well as to obtain a better understanding of mechanisms underlying major developmental events in insects, such as pupation, etc.The titer of ecdysterone in the hemolymph rises dramatically several hours before pupariation (44,56,78). This increase in ecdysterone level has long been known to induce a complex set of changes in puffing activities of polytene chromosomes (12, 14) that presumably are initiated and guided by binding of an ecdysterone-receptor complex to DNA, i.e., to promoters of regulated genes. Direct evidence for binding of ecdysterone, most likely in a complex with its receptor, to polytene chromosomes has been obtained from immunodecoration experiments with anti-ecdysterone antibodies (28, 40). Ecdysterone-induced changes in puffing activity are sequential and transient, and puffs have been labeled as early or late depending on the time of their appearance (4). The early puffs, presumably, include genes whose transcription is activated directly by ecdysterone receptor and is inhibited by their own products. Genes in late puffs may be upregulated by early gene products and downregulated by ecdysterone receptor. These interactions are described in the attractive hierarchical model of gene regulation by Ashburner and colleagues (3, 5) that, until recently, had been supported exclusively by the genetic analysis of puffing. Genes from two prominent early puffs that, pr...

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“…10 nM) and its receptor complex is kinetically unstable, success in studies of the receptor(s) has relied upon use of a more potent analog, the phytoecdysone ponasterone A (40,41), which is 10-50 times as active in biological assays and which is comparably more active as a ligand (30)(31)(32)(33)(34)(35)(36)(37)(38)42). Thus, the introduction of [3H]ponasterone A (>100 Ci/mmol; 1 Ci = 37 GBq) has been critical to recent progress in the study of ecdysone receptors (31,36,38).…”

mentioning

confidence: 99%

“…0.5 nM. Thus, at least in the Kc cell bioassay system, iodoponasterone is one of the most potent ecdysones known, exceeded in activity only by 14-desoxymuristerone (65 (1988) high-affinity binding sites for iodoponasterone are present in Kc167 cells at approximately the abundance expected for ecdysone receptors (9,(30)(31)(32)(33)(34)(35)(36)(37)(38).…”

mentioning

confidence: 99%

26-[125I]iodoponasterone A is a potent ecdysone and a sensitive radioligand for ecdysone receptors.

Cherbas

Lee

et al. 1988

Proc. Natl. Acad. Sci. U.S.A.

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The uptake and release of ponasterone a by the Kc cell line of Drosophila melanogaster

Cited by 34 publications

References 17 publications

Action of 24-epibrassinolide on a cell line of the beet armyworm,Spodoptera exigua

Action of 24-epibrassinolide on a cell line of the beet armyworm,Spodoptera exigua

Ecdysterone receptor is a sequence-specific transcription factor involved in the developmental regulation of heat shock genes.

26-[125I]iodoponasterone A is a potent ecdysone and a sensitive radioligand for ecdysone receptors.

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