The Drosophila Zinc Finger Transcription Factor Ouija Board Controls Ecdysteroid Biosynthesis through Specific Regulation of spookier

Abstract

Steroid hormones are crucial for many biological events in multicellular organisms. In insects, the principal steroid hormones are ecdysteroids, which play essential roles in regulating molting and metamorphosis. During larval and pupal development, ecdysteroids are synthesized in the prothoracic gland (PG) from dietary cholesterol via a series of hydroxylation and oxidation steps. The expression of all but one of the known ecdysteroid biosynthetic enzymes is restricted to the PG, but the transcriptional regulatory networks responsible for generating such exquisite tissue-specific regulation is only beginning to be elucidated. Here, we report identification and characterization of the C2H2-type zinc finger transcription factor Ouija board (Ouib) necessary for ecdysteroid production in the PG in the fruit fly Drosophila melanogaster. Expression of ouib is predominantly limited to the PG, and genetic null mutants of ouib result in larval developmental arrest that can be rescued by administrating an active ecdysteroid. Interestingly, ouib mutant animals exhibit a strong reduction in the expression of one ecdysteroid biosynthetic enzyme, spookier. Using a cell culture-based luciferase reporter assay, Ouib protein stimulates transcription of spok by binding to a specific ~15 bp response element in the spok PG enhancer element. Most remarkable, the developmental arrest phenotype of ouib mutants is rescued by over-expression of a functionally-equivalent paralog of spookier. These observations imply that the main biological function of Ouib is to specifically regulate spookier transcription during Drosophila development.

Altered levels ofhsromegalncRNAs further enhance Ras signaling during ectopically activated Ras induced R7 differentiation inDrosophila

“…7A, B). CG11762 and forkhead genes are part of the ecdysone signaling pathway (Cao et al, 2007;Komura-Kawa et al, 2015). Ssb-c31a, as noted earlier, is reported to bind to Raf protein.…”

The Insect Prothoracic Gland as a Model for Steroid Hormone Biosynthesis and Regulation

“…The regulatory group comprises a total of 13 DNA-binding proteins. Of these, molting defective ( mld ), vvl, ouija board ( ouib , aka CG11762 ), snail , and timeless ( tim ) were already known to have specific expression in the RG (Cheng et al, 2014; Danielsen et al, 2014; Komura-Kawa et al, 2015; Morioka et al, 2012; Neubueser et al, 2005;Sánchez-Higueras et al, 2014), while tinman has been linked to the embryonic development of the CC, but no expression in the RG has been demonstrated (Park et al, 2011). Among the identified transcription factors are Escargot, which is related to Snail, and Hand, which ranks among the 50 most highly enriched mRNAs in the RG (Table S2), as well as pdm3 , which encodes—like vvl —a POU-domain transcription factor.…”

A Drosophila Genome-Wide Screen Identifies Regulators of Steroid Hormone Production and Developmental Timing

“…Gene hits associated with arrest in L1 or L2, the strongest phenotypes, include genes directly involved in the ecdysone biosynthetic pathway ( shroud , phantom , disembodied , shadow and Cyp6t3 ). The screen also identified genes associated with cholesterol trafficking ( Npc1a , GstE14/Nobo and snmp1 ), genes in major signaling pathways such as insulin/TOR ( InR, Akt1, raptor, Tif-1A ), PTTH/Torso/Ras ( torso, Ras85D ) and TGFβ ( put and tkv ), and transcription factors ( vvl , kni , mld, ouib, br, E75B, EcR and USP ) that are known to regulate ecdysone production in the PG (Caceres et al, 2011; Colombani et al, 2005; Danielsen et al, 2014; Gibbens et al, 2011; Huang et al, 2005; Komura-Kawa et al, 2015; Koyama et al, 2014; Layalle et al, 2008; Mirth et al, 2005; Moeller et al, 2013; Niwa et al, 2010; Niwa and Niwa, 2014; Ou et al, 2011; Rewitz et al, 2009; Talamillo et al, 2013; Zhou et al, 2004). This shows that our screen was successful in identifying genes with known steroidogenic roles, and an additional ~1,800 genes that have not been linked to either steroidogenesis, steroidogenic cell function or general gland viability; indeed, many of these genes have no identified function.…”