Abstract
BackgroundThe 20-hydroxyecdysone (20E) hierarchy of gene activation serves as an attractive model system for studying the mode of steroid hormone regulated gene expression and development. Many structural analogs of 20E exist in nature and among them the plant-derived ponasterone A (PoA) is the most potent. PoA has a higher affinity for the 20E nuclear receptor, composed of the ecysone receptor (EcR) and Ultraspiracle proteins, than 20E and a comparison of the genes regulated by these hormones has not been performed. Furthermore, in Drosophila different cell types elicit different morphological responses to 20E yet the cell type specificity of the 20E transcriptional response has not been examined on a genome-wide scale. We aim to characterize the transcriptional response to 20E and PoA in Drosophila Kc cells and to 20E in salivary glands and provide a robust comparison of genes involved in each response.ResultsOur genome-wide microarray analysis of Kc167 cells treated with 20E or PoA revealed that far more genes are regulated by PoA than by 20E (256 vs 148 respectively) and that there is very little overlap between the transcriptional responses to each hormone. Interestingly, genes induced by 20E relative to PoA are enriched in functions related to development. We also find that many genes regulated by 20E in Kc167 cells are not regulated by 20E in salivary glands of wandering 3rd instar larvae and we show that 20E-induced levels of EcR isoforms EcR-RA, ER-RC, and EcR-RD/E differ between Kc cells and salivary glands suggesting a possible cause for the observed differences in 20E-regulated gene transcription between the two cell types.ConclusionsWe report significant differences in the transcriptional responses of 20E and PoA, two steroid hormones that differ by only a single hydroxyl group. We also provide evidence that suggests that PoA induced death of non-adapted insects may be related to PoA regulating different set of genes when compared to 20E. In addition, we reveal large differences between Kc cells and salivary glands with regard to their genome-wide transcriptional response to 20E and show that the level of induction of certain EcR isoforms differ between Kc cells and salivary glands. We hypothesize that the differences in the transcriptional response may in part be due to differences in the EcR isoforms present in different cell types.
Highlights
The 20-hydroxyecdysone (20E) hierarchy of gene activation serves as an attractive model system for studying the mode of steroid hormone regulated gene expression and development
Identification of 20-hydroxyecdysone (20E)-responsive genes in Kc167 cells The 20E-hierarchy of gene transcription serves as a good model for examining hormonal control of development
The response observed here after a 2 and 4 hour 20E-treatment are consistent with two prior observations on the 20E-induced puffing patterns of larval salivary glands: first, that primary 20E-response genes are likely to still be detectable after a four-hour 20Etreatment and second, that the 20E-response of Kc167 is more robust after 4 hours of 20E exposure [4]
Summary
The 20-hydroxyecdysone (20E) hierarchy of gene activation serves as an attractive model system for studying the mode of steroid hormone regulated gene expression and development. In Drosophila and other arthropods, pulses of the steroid hormone 20-hydroxyecdysone (20E) are responsible for the temporal coordination of larval molts and metamorphosis Physiological responses during these events can be diverse; for example during metamorphosis obsolete larval tissues are destroyed and adult structures of polytene chromosomes of late 3rd instar larvae in response to 20E, predicted that primary-response genes would code for proteins that are responsible both for the induction of secondary response genes as well as for the inhibition of their own transcription [1]. Three of the most well characterized 20E-primary response genes, Eip74EF, Eip75B, and br fit this description perfectly These three genes reside at chromosome cytolocations 74EF, 75B, and 2B5 respectively, which, along with approximately three other loci, exhibit rapid and dramatic puffing after exposure to 20E either naturally or artificially [2,3,4]. Since the original characterization of these 20E-response genes, many examples of primary and secondary response genes with diverse functions have emerged, and many of these 20Einducible genes do not appear to be associated with any identifiable 20E-induced puffs [7,8,9,10,11,12,13] underling the importance identifying individual components of the cascade for a clearer picture of 20E action
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