Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Transcriptional signature of progesterone-induced germinal vesicle breakdown in the fathead minnow ovary (Pimephales promelas)

ABSTRACT: A growing number of studies have examined transcriptional responses to sex steroids along the hypothalamic-pituitary-gonadal axis in teleost fishes. However, data is are lacking on the molecular signaling cascades that underlie progesterone signaling. The objectives of the present study were to characterize the transcriptional response in the ovary of fathead minnow (Pimephales promelas) (FHM) in response to progesterone (P4) using transcriptomics. Female FHM ovaries were exposed in vitro to 500 ng P4/L. Germinal vesicle migration and breakdown (GVBD) was observed and microarrays were used to identify gene cascades affected by P4. Microarray analysis identified 1,265 differentially expressed transcripts after P4 treatment (p < 0.05). Functional enrichment analysis revealed that transcripts involved in the molecular functions of protein serine/threonine kinase activity, ATP binding, and activity of calcium channels were increased in the female ovary after P4 treatment. There was an overwhelming response of genes involved in structural constituent of ribosome to bethat were down-regulated with P4. There was also evidence for gene changes in steroid and maturation-related transcripts. Pathway analyses identified cell cycle regulation, insulin action, hedgehog, and B cell activation as pathways containing an over-representation of highly regulated transcripts. Significant regulatory sub-networks of P4-mediated transcripts included genes regulated by tumor protein p53 and E2F transcription factor 1, suggesting that downstream expression targets of these two proteins may be preferentially involved in P4-mediated GVBD. These data provide novel insight into the molecular signaling cascades that may underlie P4-signaling in the ovary and identifies genes and processes that may indicate premature GVBD due to environmental pollutants that mimic progestins. 3 control and 4 treatment; biological replicates

ORGANISM(S): Pimephales promelas

SUBMITTER: Christopher Martyniuk

PROVIDER: E-GEOD-43601 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Follicle assembly is the process by which groups or ‘nests’ of oocytes break down to form primordial follicles. The size of the primordial follicle pool is the major determinant of the reproductive lifespan of a female. Previously, progesterone (P4) has been shown to inhibit follicle assembly, while tumor necrosis factor alpha (TNF-alpha) has been shown to promote the apoptosis that is necessary for follicle assembly. The current study examines how TNF-alpha and progesterone interact to regulate primordial follicle assembly. Ovaries were collected from newborn rats and placed in organ culture to examine the actions of P4 and TNF-alpha. P4 was found to decrease primordial follicle assembly and increase the percentage of un-assembled oocytes both in vitro and in vivo. TNF-alpha treatment did not change the proportion of assembled follicles in cultured ovaries, but did block the ability of P4 to inhibit follicle assembly. Microarray analysis of the ovarian transcriptome revealed that progesterone treatment of the ovaries altered the expression of 513 genes with 132 only expressed after P4 treatment and 16 only expressed in control ovaries. The majority of genes were up-regulated greater than 2-fold over control, with a small subset of 16 genes down-regulated. Categories of genes affected by P4 are described including a group of extra-cellular signaling factors. The progesterone receptors expressed at the time of follicle assembly included the surface membrane progesterone receptors PGRMC1, PGRMC2 and RDA288. The nuclear genomic P4 receptor was not expressed at appreciable levels. Progesterone increased the expression of several genes (TANK, NFkappaB, Bcl2l1 and Bcl2l2) involved in a signaling pathway that promotes cell survival and inhibits apoptosis. Observations indicate that P4 acts through the surface membrane progesterone receptors to regulate primordial follicle assembly, and that TNF-alpha can over-ride the inhibitory actions of P4 on follicle assembly. A major mechanism involved in the actions of P4 is an increase in cell survival genes and inhibition of the apoptosis pathway. Observations provide insight into the hormonal regulation of primordial follicle assembly and lead to novel approaches to potentially manipulate follicle assembly and reproductive capacity. Keywords: expression analysis, progesterone, TNFa, follicle assembly, ovary

Project description:Ovarian estrogen (E2) and progesterone (P4) are indispensable for embryo-implantation and endometrial stromal decidualization; however, the molecular mechanisms that underpin these reproductive processes are unclear. Steroid receptor coregulator-2 (SRC-2) belongs to the multifunctional SRC/p160 family which also includes SRC-1 and SRC-3. Sharing strong sequence homology, all three SRCs exert diverse regulatory effects by modulating the transcriptional potency of nuclear receptor family members, including the estrogen and progesterone receptor (ER and PR respectively). Importantly, absence of SRC-2 in PR positive cells in the epithelial, stromal, and myometrial compartments of the murine uterus results in a striking infertility defect. This reproductive phenotype highlights a key role for SRC-2 in uterine function which is not shared with other coregulators. Intriguingly, abrogation of uterine SRC-2 does not block embryo apposition or attachment to the apical surface of luminal epithelial cells of the endometrium but rather prevents P4-dependent local decidualization of the sub-epithelial stroma. Remarkably, epithelial-specific ablation of SRC-2 in the murine uterus does not compromise endometrial functionality, again underscoring the unique importance of stromal derived SRC-2 in uterine function. The stromal decidualization defect resulting from SRC-2 ablation is reflected at the molecular level by a marked attenuation in P4 responsive target genes known to be critical for P4 dependent decidualization (i.e. ERBB receptor feedback inhibitor 1, Follistatin and Fkbp5). Conversely, the induction of E2 or P4 target genes involved in embryo implantation (i.e. leukemia inhibitory factor (LIF) and Indian hedgehog (Ihh) respectively) is not affected by SRC-2’s absence. As with mouse studies, decidualization of primary human stromal cells (HESCs) in culture is blocked by SRC-2 knockdown; however, HESC decidualization is unaffected by knockdown of SRC-1 or SRC-3. As a consequence of SRC-2 knockdown, molecular studies disclose a striking decrease in the induction of a subset of P4 target genes (i.e. WNT4 and FKBP5) which are essential for the stromal-epithelioid transformation step, the cellular hallmark of endometrial decidualization. Collectively, these studies not only showcase the evolutionary importance of SRC-2 in endometrial biology but also suggest that deregulation of this coregulator may underpin a spectrum of hormone-dependent uterine pathologies such as endometriosis and endometrial cancer. Microarray analysis was performed on mouse uteri using eighteen SRC-2flox/flox (SRC-2f/f) and eighteen PRCre/+ SRC-2flox/flox (SRC-2d/d) mice. Mice were ovariectomized at 6 weeks and after 2 weeks mice were either treated with sesame oil (vehicle) or 1 mg of P4. RNA from three mice per genotype per treatment were pooled and assigned as one sample (three samples per genotype per treatment). multiple group comparison

Project description:Follicle assembly is the process by which groups or ânestsâ of oocytes break down to form primordial follicles. The size of the primordial follicle pool is the major determinant of the reproductive lifespan of a female. Previously, progesterone (P4) has been shown to inhibit follicle assembly, while tumor necrosis factor alpha (TNF-alpha) has been shown to promote the apoptosis that is necessary for follicle assembly. The current study examines how TNF-alpha and progesterone interact to regulate primordial follicle assembly. Ovaries were collected from newborn rats and placed in organ culture to examine the actions of P4 and TNF-alpha. P4 was found to decrease primordial follicle assembly and increase the percentage of un-assembled oocytes both in vitro and in vivo. TNF-alpha treatment did not change the proportion of assembled follicles in cultured ovaries, but did block the ability of P4 to inhibit follicle assembly. Microarray analysis of the ovarian transcriptome revealed that progesterone treatment of the ovaries altered the expression of 513 genes with 132 only expressed after P4 treatment and 16 only expressed in control ovaries. The majority of genes were up-regulated greater than 2-fold over control, with a small subset of 16 genes down-regulated. Categories of genes affected by P4 are described including a group of extra-cellular signaling factors. The progesterone receptors expressed at the time of follicle assembly included the surface membrane progesterone receptors PGRMC1, PGRMC2 and RDA288. The nuclear genomic P4 receptor was not expressed at appreciable levels. Progesterone increased the expression of several genes (TANK, NFkappaB, Bcl2l1 and Bcl2l2) involved in a signaling pathway that promotes cell survival and inhibits apoptosis. Observations indicate that P4 acts through the surface membrane progesterone receptors to regulate primordial follicle assembly, and that TNF-alpha can over-ride the inhibitory actions of P4 on follicle assembly. A major mechanism involved in the actions of P4 is an increase in cell survival genes and inhibition of the apoptosis pathway. Observations provide insight into the hormonal regulation of primordial follicle assembly and lead to novel approaches to potentially manipulate follicle assembly and reproductive capacity. Experiment Overall Design: RNA samples from two control groups (pooled untreated cultured ovaries) are compared to two treated groups (pooled cultured ovaries treated with progesterone)

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Transcriptional signature of progesterone-induced germinal vesicle breakdown in the fathead minnow ovary (Pimephales promelas)

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