The Sf1-related nuclear hormone receptor Hr39 regulates Drosophila female reproductive tract development and function
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ABSTRACT: The vertebrate nuclear hormone receptor steroidogenic factor 1 (SF1; NR5A1) controls reproductive development and regulates the transcription of steroid-modifying cytochrome P450 genes. We find that the SF1-related Drosophila nuclear hormone receptor HR39 is also essential for sexual development. In Hr39 mutant females, the sperm-storing spermathecae and glandular parovaria are absent or defective, causing sterility. Our results indicate that spermathecae and parovaria secrete reproductive tract proteins required for sperm maturation and function, like the mammalian epididymis and female reproductive tract. Hr39 controls the expression of specific cytochrome P450 genes and is required in females both to activate spermathecal secretion and repress male-specific courtship genes such as takeout. Thus, a pathway that, in vertebrates, controls sex-specific steroid hormone production, also mediates reproductive functions in an invertebrate. Our findings suggest that Drosophila can be used to model more aspects of mammalian reproductive biology than previously believed. Keywords: mutant/wild-type comparison and tissue comparison
Project description:The vertebrate nuclear hormone receptor steroidogenic factor 1 (SF1; NR5A1) controls reproductive development and regulates the transcription of steroid-modifying cytochrome P450 genes. We find that the SF1-related Drosophila nuclear hormone receptor HR39 is also essential for sexual development. In Hr39 mutant females, the sperm-storing spermathecae and glandular parovaria are absent or defective, causing sterility. Our results indicate that spermathecae and parovaria secrete reproductive tract proteins required for sperm maturation and function, like the mammalian epididymis and female reproductive tract. Hr39 controls the expression of specific cytochrome P450 genes and is required in females both to activate spermathecal secretion and repress male-specific courtship genes such as takeout. Thus, a pathway that, in vertebrates, controls sex-specific steroid hormone production, also mediates reproductive functions in an invertebrate. Our findings suggest that Drosophila can be used to model more aspects of mammalian reproductive biology than previously believed. Experiment Overall Design: Wild type and Hr39(04443) Spermathecae, Wild type and Hr39(04443) Reproductive Tract
Project description:We use closely-related Drosophila species to understand mechanosensation and how hearing as a form of mechanosensation drives reproductive isolation and evolution. In particular, we performed RNA-seq to obtain the Johnston's Organ transcriptomes of six closely-related Drosophila species, namely, D.melanogaster, D.yakuba, D.pseudoobscura and D.persimilis. Application of the Ornstein-Uhlenbeck model identified gene expression changes between the species' auditory structures. Motif discovery ( i-cisTarget and iRegulon) in differentially expressed genes in the oobscura group and melanogaster group on these transcriptomes identified thetranscription factor Hr39 as important for auditory differences among species. Hr39 is a nuclear hormone receptor important in song production and spermathecae development. Futher functional analyses Hr39 and many of its downstream targets are expressed in Johnston's Organ neurons. Hr39 mutants show sexually dimorphic defects in auditory mechanics and compound action potentials. These results indicate that Hr39 and its downstream regulon have a key role in Drosophila courtship behavior as a mechanism of reproductive isolation and evolution.
Project description:The role of estrogen and testosterone in the regulation of gene expression in the proximal reproductive tract is not completely understood. To address this question, mice were treated with testosterone or estradiol and RNA from the efferent ducts and caput epididymis was processed and hybridized to Affymetrix MOE 430 2.0 microarrays. Analysis of array output identified probe sets in each tissue with altered levels in hormone treated versus control animals. Hormone treatment efficacy was confirmed by determination of serum hormone levels pre- and post-treatment and observed changes in transcript levels of previously reported hormone-responsive genes. Tissue-specific hormone sensitivity was observed with 2867 and 3197 probe sets changing significantly in the efferent ducts after estrogen and testosterone treatment, respectively. In the caput epididymis, 117 and 268 probe sets changed after estrogen and testosterone treatment, respectively, demonstrating a greater response to hormone in the efferent ducts than the caput epididymis. Transcripts sharing similar profiles in the intact and hormone-treated animals compared with castrated controls were also identified. Ontological analysis of probe sets revealed a significant number of hormone-regulated transcripts encode proteins associated with lipid metabolism, transcription and steroid metabolism in both tissues. Real-time RT-PCR was employed to confirm array data and investigate other potential hormone-responsive regulators of proximal reproductive tract function. The results of this work reveal previously unknown responses to estrogen in the caput epididymis and to testosterone in the efferent ducts as well as tissue specific hormone sensitivity in the proximal reproductive tract. Adult animals were castrated or sham-castrated, allowed to recover for 14 days, and then treated with 0.015 mg estradiol (castrated), 0.015 mg testosterone propionate (castrated), or vehicle (castrated and sham-castrated as biological controls) in duplicate. Efferent duct and caput epididymis was collected from each sample and analyzed. Duplicates are included in the provided data and numbered 1 or 2 for each treatment regimen.
Project description:The role of estrogen and testosterone in the regulation of gene expression in the proximal reproductive tract is not completely understood. To address this question, mice were treated with testosterone or estradiol and RNA from the efferent ducts and caput epididymis was processed and hybridized to Affymetrix MOE 430 2.0 microarrays. Analysis of array output identified probe sets in each tissue with altered levels in hormone treated versus control animals. Hormone treatment efficacy was confirmed by determination of serum hormone levels pre- and post-treatment and observed changes in transcript levels of previously reported hormone-responsive genes. Tissue-specific hormone sensitivity was observed with 2867 and 3197 probe sets changing significantly in the efferent ducts after estrogen and testosterone treatment, respectively. In the caput epididymis, 117 and 268 probe sets changed after estrogen and testosterone treatment, respectively, demonstrating a greater response to hormone in the efferent ducts than the caput epididymis. Transcripts sharing similar profiles in the intact and hormone-treated animals compared with castrated controls were also identified. Ontological analysis of probe sets revealed a significant number of hormone-regulated transcripts encode proteins associated with lipid metabolism, transcription and steroid metabolism in both tissues. Real-time RT-PCR was employed to confirm array data and investigate other potential hormone-responsive regulators of proximal reproductive tract function. The results of this work reveal previously unknown responses to estrogen in the caput epididymis and to testosterone in the efferent ducts as well as tissue specific hormone sensitivity in the proximal reproductive tract.
Project description:Male Anophele gambiae mosquitoes transfer the steroid hormone 20-hydroxyecdysone (20E) to the female lower reproductive tract during mating. We have shown that this sexually transferred hormone is a key regulator of monandry and oviposition in An. gambiae females. In order to identify possible molecular pathways underlying the effects induced by sexually transferred 20E we investigated the transcriptional response in the atrium and spermatheca at 24 hours after 20E injection in the thorax of virgin females, which delivers to these reproductive tissues 20E levels comparable to those detected after mating. By comparing these results to those of microarrays assessing the post-mating transcriptional response in the female lower reproductive tract (LRT) at 24 hours post copulation, we were able to identify mating responsive genes likely regulated by male transferred 20E. For each experiment atrium and spermatheca were dissected from females 24 hours after 20E or ethanol (10%) injection and transcriptional profiles compared across 4 biological replicates using Agilent two-color microarrays
Project description:We compared gene expression in oviduct tissues between unmated (control) and mated hen. As spermatozoa are foreign to the female reproductive tract therefore we were interested to look at how spermatozoa survive in the female reproductive tract and keep their fertilization potentiality. To check that we collected tissues from oviduct of both control and mated female chicken and compared if sperm deposition to the oviduct made any gene expression shift related to sperm survival.
Project description:The HIF (hypoxia-inducible factor) transcription factor is the master regulator of the metazoan response to chronic hypoxia. In addition to promoting adaptations to low oxygen, HIF drives cytoprotective mechanisms in response to stresses and modulates neural circuit function. How most HIF targets act in the control of the diverse aspects of HIF-regulated biology remains unknown. We discovered that a HIF target, the C. elegans gene cyp-36A1, is required for numerous HIF-dependent processes, including modulation of gene expression, stress resistance, and behavior. cyp-36A1 encodes a cytochrome P450 enzyme that we show controls expression of more than a third of HIF-induced genes. CYP-36A1 acts cell non-autonomously by regulating the activity of the nuclear hormone receptor NHR-46, suggesting that CYP-36A1 functions as a biosynthetic enzyme for a hormone ligand of this receptor. We propose that regulation of HIF effectors through activation of cytochrome P450 enzyme/nuclear receptor signaling pathways could similarly occur in humans.
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: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. Keywords: keywords: reproduction, reproductive tract, accessory gland proteins, sperm, timecourse