Project description:Chromatin modifications have essential roles in directing nervous system development and behavior. Drosophila courtship is an ideal model for understanding the relationships between chromatin modifications, gene expression, sexual dimorphisms in neuroanatomy, and reproductive behaviors. These behaviors are largely innate and are under the control of sex determination hierarchy genes doublesex (dsx) and fruitless (fru) that encode sex-specific transcription factors. Transcripts from the fru P1 promoter are sex-specifically spliced, resulting in male-specific proteins, FruM, which are required for male courtship behaviors. FruM has been shown to form a complex with Bonus/TIF1, a transcriptional cofactor, and interact with chromatin modifying proteins. This indicates that sex-differential modification of the chromatin landscape contributes to creating and maintaining the potential for sexually dimorphic behavior. To examine chromatin modifications genome-wide in fru P1-expressing neurons, we have developed a method to purify chromatin in a cell-type-specific manner using a tagged histone H2B under UAS control, “Chromatag”, paired with sequential ChIP-seq. Here, we evaluate five chromatin modifications: H3K27ac, H3K4me3, H3K36me3, H3K9me3, and H3K27me3 to investigate fru P1-expressing neurons at three developmental stages in both sexes. These modifications have been well characterized for their roles in transcriptional activation or repression. We observe greater changes in chromatin modification profiles across developmental stages than between sexes. In addition, we have generated cell-type specific RNA-seq data sets, using Translating Ribosome Affinity Purification (TRAP), to examine gene expression in fru P1-expressing neurons of both sexes. Informed by these data sets, we have identified several genes with roles in neuronal function and examined their expression in fru P1-expressing neurons, revealing sexually dimorphic expression patterns. Altogether, this work offers insights into stage and sex-differences in chromatin modifications and gene expression in fru P1-expressing neurons.

Project description:Propranolol is a beta-adrenergic receptor antagonist (Î²-blocker) that has been detected in United States wastewater effluents at concentrations ranging from 0.026 to 1.90 Âµg/l. In mammals, there is evidence that Î²-blockers can cause sexual dysfunction, and alter serotonergic pathways which may impact reproductive behavior but little is known about the effects on fish behavior. The present study tested the effects of propranolol on fecundity and on reproductive behavior of the fathead minnow, Pimephales promelas, a fish that exhibits male parental care. Sexually mature fathead minnows were housed at a ratio of one male and two females per tank and exposed to nominal concentrations of 0, 0.1, 1, 10 Âµg/l for 21 days. Measured concentrations (Â±SD) of propranolol were 0.05Â±0.02, 0.88Â±0.34 and 4.11Â±1.19 Âµg/l. There were no statistically significant differences in fecundity, fertilization rate, hatchability and time to hatch. Propranolol exposure was not associated with a change in nest rubbing behavior, time spent in the nest or approaching the females. There was a significant difference in the number of visits to the nest with males receiving low and medium propranolol treatments. The microarray analysis showed that there were 335 genes up-regulated and 400 genes down-regulated in the brain after exposure to the highest dose of propranolol. Among those genes, myoglobin and calsequestrin transcripts (fold change=10.84 and 5.49, respectively) were highly up-regulated. Ontological analyses indicated changes in genes involved in calcium ion transport, transcription, proteolysis and apoptosis/anti-apoptosis. The results showed that exposure to propranolol at concentrations as high as 4.11 Âµg/l did not significantly impact reproductive behavior or spawning abilities of fathead minnow but did alter the regulation of genes within the brain of fish. Effects of propanolol exposure were investigated in the brain of adult male fathead minnow exposed to 10 Âµg/L of propanolol or a solvent control solution (0.01% ethanol). For each treatment, the brain of four different fish were analyzed.

Project description:Adult reproductive behaviors in Drosophila melanogaster males and females are vastly different. Yet, neurons that express sex-specifically spliced fruitless transcripts (fru P1) underlie these behaviors in both sexes. How the same set of neurons can drive such different behaviors is an important and unresolved question in developmental genetics. A particular challenge is that fru P1-expressing neurons are only 2-5% of the adult nervous system, making studies of adult head tissue, or even the whole brain, unlikely to yield informative inferences. Translating Ribosome Affinity Purification (TRAP) identifies the actively translated pool of mRNAs from fru P1-expressing neurons and allowed us to conduct a sensitive, cell-specific assay of gene expression. The male and female fru P1-expressing neurons have a shared set of 1,642 genes with enriched TRAP transcripts that form a distinct repertoire, relative to TRAP analyses of all neurons in the adult head. Further, there are a striking number of genes (3,147) that have sex-biased TRAP enrichment in fru P1-expressing neurons. Yet, most of these genes (3,107) have only male-biased TRAP enrichment. This suggests an underlying mechanism to generate dimorphism in behavior, with the transcript repertoire that specifies female behaviors present in both sexes and a large additional set of genes with expression in the male dramatically altering the pattern. Thus, these additional genes invoke the male-specific behaviors by establishing cell fate in the same context of gene expression observed in females. These results suggest a possible global mechanism for how distinct behaviors can arise in different environments, from a shared set of neurons. Libraries were prepared from five independent biological replicates, from TRAP adult heads samples from males and females, and the mRNA input from adult heads from males and females. For each experimental condition, approximately 1,000 flies that were 8 to 24 hours post-eclosion were used.

Dataset Information

Analysis of cell-type-specific chromatin modifications and gene expression in Drosophila neurons that direct reproductive behavior [TRAP I]

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