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Drosophila E93 promotes adult development and suppresses larval responses to ecdysone during metamorphosis

ABSTRACT: Pulses of the steroid hormone ecdysone act through transcriptional cascades to direct the major developmental transitions during the Drosophila life cycle. These include the prepupal ecdysone pulse, which occurs 10 hours after pupariation and triggers the onset of adult morphogenesis and larval tissue destruction. E93 encodes a transcription factor that is specifically induced by the prepupal pulse of ecdysone, supporting a model proposed by earlier work that it specifies the onset of adult development. Although a number of studies have addressed these functions for E93, little is known about its roles in the salivary gland where the E93 locus was originally identified. Here we show that E93 is required for development through late pupal stages, with mutants displaying defects in adult differentiation and no detectable effect on the destruction of salivary glands. RNA-seq analysis demonstrates that E93 regulates genes involved in development and morphogenesis in the salivary glands, but has little effect on cell death gene expression. We also show that E93 is required to direct the proper timing of ecdysone-regulated gene expression in salivary glands, and that it suppresses earlier transcriptional programs that occur during larval and prepupal stages. These studies support the model that the stage-specific induction of E93 in late prepupae provides a critical signal that defines the end of larval development and the onset of adult differentiation.

ORGANISM(S): Drosophila melanogaster

PROVIDER: GSE181948 | GEO | 2022/08/11

REPOSITORIES: GEO

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Project description:Although the salivary transcriptome of adult mosquitoes has been thoroughly described in several recent papers, very little information exists regarding the biological role of larval salivary glands in the Culicidae. We used whole-transcriptome AffymetrixÂ® chips to compare the transcriptional profiles of Anopheles gambiae larval (L4) salivary glands and whole larvae. We identified a total of 277 transcripts as being significantly enriched in the salivary glands. Based on available annotation for the known or predicted protein sequences encoded by these transcripts, 41 were identified as corresponding to secreted proteins, 233 as corresponding to non-secreted (housekeeping) proteins, and 3 as encoding proteins of unknown function. Based on functional annotation of the putatively secreted gene products, we propose that larval salivary secretions have roles in nutrient digestion, detoxification, immunity, and mouthpart lubrication. Interestingly, several components of the larval saliva (e.g. apyrase and serine proteases) have also been reported to exist in adult female saliva, where they are though to help regulate a vertebrate hostâs immune response to bloodfeeding. In conclusion, our results suggest that the salivary glands are important components of both the digestive and immune systems of larval mosquitoes, and that their study might provide clues about the evolution of adaptations to bloodfeeding observed in adults. Experiment Overall Design: We attempted to identify transcripts that are both present and significantly enriched in the salivary glands (SG) of the 4th instar Anopheles gambiae larva. Transcripts were considered as present in the SG if the built-in Affymetrix detection call scored them as such in all three biological replicates of the experiment. Expression values (i.e. signal intensities) of transcripts from the salivary glands were compared to those of a matching group of whole 4th instar larvae using a t-test. Transcripts showing an expression value two-fold or higher (p < 0.01; false discovery rate=1%) in the SG as compared to whole larvae were considered to be significantly enriched.

Project description:Analysis of differential gene expression in third instar Drosophila salivary glands in the absence versus presence of cohesin. ABSTRACT: Developmental abnormalities observed in Cornelia de Lange Syndrome (CdLS) have been genetically linked to mutations in the cohesin machinery. These findings raise the possibility that cohesin, in addition to its canonical function of mediating sister chromatid cohesion, might also be involved in regulating gene expression. We report that cleavage of cohesinM-CM-^Us kleisin subunit in post-mitotic Drosophila salivary glands induces major changes (both up and down) in the transcript levels of many genes. Kinetic analyses of changes in transcript levels upon cohesin cleavage reveal that a subset of genes responds to cohesin cleavage within a few hours. In addition, cohesin binds to most of these loci, suggesting that cohesin is directly regulating their expression. Amongst these genes are several that are regulated by the steroid hormone Ecdysone. Transcripts at EcR and Eip74EF, which encode an Ecdysone Receptor and an Ecdysone-regulated transcription factor, respectively, decline ten-fold within four hours of cohesin cleavage. Cytological visualization of transcription at selected Ecdysone-responsive genes reveals that puffing at Eip74EF ceases within an hour or two of cohesin cleavage, long before any decline in EcR associated with this locus. We conclude that cohesin regulates expression of a distinct set of genes, including those mediating the Ecdysone response. A heat-inducible transgene (hs-TEV) was used to induce TEV in terminally differentiated third instar Drosophila salivary glands expressing either wild type (+ cohesin) or TEV-cleavable myc10-tagged Rad21 protein (Rad21TEV, - cohesin). Total RNA was isolated from + and - cohesin salivary glands 10-12 hours after heat shock induction of TEV (7 independent biological samples each). RNA samples were converted to cDNA, labeled with Cy3 and Cy5 respectively (3x) and vice versa (4x; dye swaps), and hybridized to INDAC FL003 arrays. Analysis of seven arrays, each hybridized to an independently generated sample-pair revealed major differences in transcript levels between + and - cohesin samples.

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Drosophila E93 promotes adult development and suppresses larval responses to ecdysone during metamorphosis

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