Regulation of Drosophila Metamorphosis by Xenobiotic Response Regulators

University of California San Francisco, United States of America
PLoS Genetics (Impact Factor: 7.53). 02/2013; 9(2):e1003263. DOI: 10.1371/journal.pgen.1003263
Source: PubMed


Mammalian Nrf2-Keap1 and the homologous Drosophila CncC-dKeap1 protein complexes regulate both transcriptional responses to xenobiotic compounds as well as native cellular and developmental processes. The relationships between the functions of these proteins in xenobiotic responses and in development were unknown. We investigated the genes regulated by CncC and dKeap1 during development and the signal transduction pathways that modulate their functions. CncC and dKeap1 were enriched within the nuclei in many tissues, in contrast to the reported cytoplasmic localization of Keap1 and Nrf2 in cultured mammalian cells. CncC and dKeap1 occupied ecdysone-regulated early puffs on polytene chromosomes. Depletion of either CncC or dKeap1 in salivary glands selectively reduced early puff gene transcription. CncC and dKeap1 depletion in the prothoracic gland as well as cncC(K6/K6) and dKeap1(EY5/EY5) loss of function mutations in embryos reduced ecdysone-biosynthetic gene transcription. In contrast, dKeap1 depletion and the dKeap1(EY5/EY5) loss of function mutation enhanced xenobiotic response gene transcription in larvae and embryos, respectively. Depletion of CncC or dKeap1 in the prothoracic gland delayed pupation by decreasing larval ecdysteroid levels. CncC depletion suppressed the premature pupation and developmental arrest caused by constitutive Ras signaling in the prothoracic gland; conversely, constitutive Ras signaling altered the loci occupied by CncC on polytene chromosomes and activated transcription of genes at these loci. The effects of CncC and dKeap1 on both ecdysone-biosynthetic and ecdysone-regulated gene transcription, and the roles of CncC in Ras signaling in the prothoracic gland, establish the functions of these proteins in the neuroendocrine axis that coordinates insect metamorphosis.

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    • "2B, 74EF and 75B). These puffs were bound prominently both by endogenous dKeap1 and CncC as well as by independently expressed dKeap1 and CncC fusion proteins (Fig. 1B, Fig. 2A,C; supplementary material Fig. S2A) (Deng and Kerppola, 2013). The BiFC complexes bound to fewer loci than either dKeap1 or CncC that were expressed separately, suggesting that dKeap1-CncC complexes bound chromatin with a higher specificity than either dKeap1 or CncC individually (Fig. 2C). "
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    ABSTRACT: Interactions among transcription factors control their physiological functions by regulating their binding specificities and transcriptional activities. We implement a strategy to visualize directly the genomic loci that are bound by multi-protein complexes in single cells in Drosophila. This method is based on bimolecular fluorescence complementation (BiFC) analysis of protein interactions on polytene chromosomes. Drosophila Keap1 (dKeap1)-CncC complexes localized to the nucleus and bound chromatin loci that were not bound preferentially by dKeap1 or CncC when they were expressed separately. dKeap1 and CncC binding at these loci was enhanced by phenobarbital, but not by tert-butylhydroquinone (tBHQ) or paraquat. Endogenous dKeap1 and CncC activated transcription of the Jheh (Jheh1, Jheh2, Jheh3) and dKeap1 genes at these loci, whereas CncC alone activated other xenobiotic response genes. Ectopic dKeap1 expression increased CncC binding at the Jheh and dKeap1 gene loci and activated their transcription, whereas dKeap1 inhibited CncC binding at other xenobiotic response gene loci and suppressed their transcription. The combinatorial chromatin-binding specificities and transcriptional activities of dKeap1-CncC complexes mediated the selective activation of different sets of genes by different xenobiotic compounds, in part through feed-forward activation of dKeap1 transcription.
    Development 07/2014; 141(16). DOI:10.1242/dev.110528 · 6.46 Impact Factor
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    ABSTRACT: Xenobiotic and oxidative responses protect cells from external and internal toxicities. Nrf2 and Keap1 are central factors that mediate these responses, and are closely related with many human diseases. In a recent study, we revealed novel developmental function and regulatory mechanism of Nrf2 and Keap1 by investigating their Drosophila homolog CncC and dKeap1. We found that CncC and dKeap1 control metamorphosis through regulations of ecdysone biosynthetic genes and ecdysone response genes in different tissues. CncC and dKeap1 cooperatively activate these developmental genes, in contrast to their conserved antagonizing effect to xenobiotic response transcription. In addition, interactions between CncC and Ras signaling in metamorphosis and in transcriptional regulation were established. Here I discuss the implications that place these classic xenobiotic response factors into a broader network that potentially control development and oncogenesis using mechanisms other than those mediating xenobiotic response.
    Fly 11/2013; 8(1). DOI:10.4161/fly.27007 · 3.33 Impact Factor
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    ABSTRACT: 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 βFTZ-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.
    Developmental Biology 01/2014; 387(2). DOI:10.1016/j.ydbio.2014.01.006 · 3.55 Impact Factor
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