PATHWAYS BY WHICH DROSOPHILA MELANOGASTER SEMINAL FLUID PROTEINS INFLUENCE FEMALE MIDGUT PHYSIOLOGY AND OVULATION
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In many species, seminal fluid proteins (Sfps) are necessary for both male and female reproductive success. In Drosophila melanogaster, these proteins trigger a suite of female behavioral and physiological changes collectively referred to as the post-mating response (PMR). Many of the changes in the PMR increase female reproductive output. These changes include increasing rates of oogenesis and ovulation, as well as changing food intake and midgut physiology to meet the nutritional demands of egg production. However, despite the important reproductive roles of Sfps, the molecular pathways within the female that link Sfps to PMR phenotypes remain poorly characterized. As a result, I undertook two projects focused on identifying how the Sfps sex peptide (SP) and ovulin influence aspects of the female PMR. First, I showed that SP is both necessary and sufficient for female midgut growth. Post-mating gut growth is an important part of the female PMR, as it is thought to help nutritionally support increased egg production. I then showed that SP’s role in post-mating gut growth occurs at least partially through its receptor, SPR. Moreover, RNA-seq analysis revealed that SP is responsible for nearly all midgut transcriptomic changes that are seen following mating, including up-regulation of protein and lipid metabolism genes and down-regulation of carbohydrate metabolism genes. These changes in midgut gene expression likely help supply the female with the nutrients required to sustain egg production. Thus, we report a novel role for SP in altering female physiology to enhance reproductive output: SP triggers the switch from a virgin to a mated state within the midgut. My second project focused on identifying the female receptor for ovulin, an Sfp that increases the activity of octopaminergic neurons within the female reproductive tract to stimulate ovulation in the first 24 hours after mating. My collaborators and I used an evolutionary rate co-variation screen to identify GPCRs which co-evolve with ovulin. We subsequently screened strong candidates for ovulation defects upon knockdown, as well as for genetic interactions with ovulin. These studies identified a novel role for three receptors (SIFaR, Lgr3, GabaβR1): they are needed to increase ovulation at early post-mating timepoints. By tissue-specific knockdown I then showed that SIFaR and Lgr3 expression in octopaminergic neurons is required for normal post-mating increases in ovulation. Lgr3 and GabaβR1 show phenotypes consistent with ovulin receptor activity, and will be tested for physical interactions with ovulin. Overall, this work identifies promising ovulin receptor candidates, and also expands our understanding of the signaling pathways controlling early post-mating ovulation events. Through these two projects, I have made significant progress in determining the mechanisms by which two Sfps trigger female post-mating responses.
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Supplemental file(s) description: Molecular Function GO Analysis for Midgut RNA-seq, Biological Process GO Analysis for Midgut RNA-seq, Differential Expression Data for Midgut RNA-seq.
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