Project description:Male-derived accessory gland proteins (Acps) that are transferred to females during mating have profound effects on female reproductive physiology including increased ovulation, mating inhibition, and effects on sperm utilization and storage. The extreme rates of evolution seen in Acps may be driven by sperm competition and sexual conflict, processes which may ultimately drive complex interactions between female- and male-derived molecules and sperm. However, little is known of how gene expression in female reproductive tissues changes in response to the presence of male molecules and sperm. To characterize this response, we conducted parallel genomic and proteomic analyses of gene expression in the reproductive tract of 3-day-old unmated and mated female Drosophila melanogaster. Using DNA microarrays, we identified 539 transcripts that are differentially expressed in unmated vs. mated females and revealed a striking peak in differential expression at 6 hrs postmating and a marked shift from primarily down-regulated to primarily up-regulated transcripts within 3 hrs after mating. Combining two-dimensional gel electrophoresis and liquid chromatography mass spectrometry analyses, we identified 84 differentially expressed proteins at 3 hrs postmating, including proteins which appeared to undergo post-translational modification. Together, our observations define transcriptional and translational response to mating within the female reproductive tract and suggest a bimodal model of postmating gene expression initially correlated with mating and the final stages of female reproductive tract maturation and later with the declining presence of male reproductive molecules and with sperm maintenance and utilization. Experiment Overall Design: Three-day-old mated and unmated females were dissected to remove the lower reproductive tract (upper uterus, sperm-storage organs, and accessory glands). Mated females were dissected either immediately following mating (0 hr) or at 3, 6, or 24 hrs following the termination of mating. Tracts of 12-40 females of like category were pooled and total RNA extracted via a TRIzol-based protocol. Processing and labeling of transcript was performed by the Molecular Biology Core Facility at the Medical College of Georgia. Arrays from mated females at the different timepoints were compared to unmated females.
Project description:The female’s reproductive tract is exposed directly to the male’s ejaculate, making it a hotspot for mating-induced responses shortly after mating. In Drosophila melanogaster, changes in the reproductive tract are essential to optimize fertilization. To detect the earliest gene regulatory events that underlie these changes, we measured transcript abundances using RNA-seq and microRNA-seq of reproductive tracts of unmated females and females collected within 10-15 minutes after the end of mating, either to a wildtype male or to a male with defective BMP signaling in secondary cells of the accessory gland, which influences the composition of the male’s ejaculate. We observed transcript abundance changes for genes with roles in tissue morphogenesis, wound healing, the immune response and metabolism. Strikingly, predicted targets of microRNAs that respond to mating are enriched for overlapping functions, suggesting that mating-induced changes are in part regulated by microRNAs. Most of the differentially expressed RNAs are upregulated in response to mating, while most of the differentially expressed microRNAs are downregulated. This pattern suggests a response of activation and de-repression of gene programs that switch the reproductive tract to a “mated” state, rather than a repression of virgin-specific programs. Male genotype did not influence transcript levels, indicating that the earliest transcriptomic responses in the reproductive tract are not dependent on ejaculate components that require BMP signaling in secondary cells. Our results shed light on the molecular changes that accompany very early responses to mating and present candidate genes and microRNAs that can be further examined for their participation in alterations of the reproductive tract microenvironment in response to signals from the male.
Project description:Since Anopheles gambiae female mosquitoes mate only once in their lifetime, and store sperm received from males potentially for weeks, genes and pathways regulated by mating in the sperm storage organ (spermatheca) to preserve sperm function may represent new targets for vector control. The transcriptional response to mating in the spermatheca was investigated using Agilent two-color microarrays. Spermathecae from mated females were collected 24 h after mating and compared to age-matched virgins. 4 biological replicates were performed.
Project description:Temporal gene expression changes in head tissues from mated females was determined at four time points post-mating, from 0-2 hours out to 72 hours. Each time point assayed post-mating showed a unique post-mating gene expression response, with 48 hours post-mating having the largest number of genes with expression changes. At most time points, a marked change in expression of genes expressed in the head fat body and encode products that function in metabolism was observed. Additionally, gene expression was analyzed at 24 hours post-mating in brain tissues, identifying the repressed expression of several genes encoding ion channels.
Project description:Purpose: Mating induces a multitude of changes in female behavior, physiology and gene expression. Interactions between female and male genotype lead to variation in post-mating phenotypes and reproductive success. So far, few female molecules responsible for these interactions have been identified. Methods: We used Drosophila melanogaster from five geographically dispersed populations to investigate such female x male genotypic interactions at the female transcriptomic and phenotypic levels. Methods: Females from each line were singly-mated to males from the same five lines, for a total of 25 combinations. To assess whether female x male genotypic interactions affect the female post-mating transcriptome, next-generation RNA sequencing was performed on virgin and mated females at 5 to 6 hours post-mating. Results: Seventy-seven genes showed strong variation in mating-induced expression changes in a female x male genotype-dependent manner. These genes were enriched for immune response and odorant-binding functions, and for expression exclusively in the head. Conclusions: The transcriptional variation found in specific functional classes of genes might be a read-out of female x male compatibility at a molecular level. Understanding the roles these genes play in the female post-mating response will be crucial to better understand the evolution of post-mating responses and related conflicts between the sexes.