Biology subjects

Wolfe, M. W.

Publications and source records attributed to Wolfe, M. W..

6 recordsLinked to original sources

bioRxiv2021

ESR2 regulates indian hedgehog signaling in neonatal rat ovary

The transcriptional regulatory function of estrogen receptor {beta} (ESR2) is essential for the regulation of primordial follicle activation (PFA). Increased PFA due to the loss of ESR2 becomes evident as early as postnatal day 8 (PND8). To identify the ESR2-regulated genes that control PFA, we performed RNA-seq analyses of wildtype, and Esr2 knockout (Esr2KO) neonatal rat ovaries collected on PND4, PND6, and PND8. Among the differentially expressed genes in Esr2KO ovaries, indian hedgehog (Ihh) displayed the highest downregulation among the ovary enriched genes. IHH regulated genes including Hhip as well as the steroidogenic enzymes were also downregulated in Esr2KO rat ovaries. Remarkably, the expression of Ihh in Esr2KO ovaries was not upregulated despite the high levels of Gdf9 and Bmp15, which are known regulators of Ihh expression in granulosa cells. Our findings suggest that indian hedgehog signaling in the neonatal rat ovary is dependent on ESR2.

developmental biology↗

bioRxiv2020

Granulosa cell genes that regulate ovarian follicle development beyond the antral stage: the role of estrogen receptor β

Follicle development beyond the preantral stage is dependent on gonadotropins. FSH signaling is crucial for the advancement of preantral follicles to the antral stage, and LH signaling is essential for further maturation of preovulatory follicles. Estrogen is intricately tied to gonadotropin signaling during the advanced stages of folliculogenesis. We observed that Er{beta}null ovarian follicles fail to develop beyond the antral stage, even after exogenous gonadotropin stimulation. As ER{beta} is primarily expressed in the granulosa cells (GCs), we explored the gonadotropin-regulated GC genes that induce maturation of antral follicles. Synchronized follicle development was induced by administration of exogenous gonadotropins to wildtype 4-wk-old female rats. The GC transcriptome was analyzed via RNA-sequencing before and after gonadotropin stimulation. An Er{beta}null mutant model that fails to show follicle maturation was also included in order to identify the ER{beta}-regulated genes involved at this step. We observed that specific groups of genes were differentially expressed in response to PMSG or hCG administration in wildtype rats. While some of the PMSG or hCG-induced genes showed a similar expression pattern in Er{beta}null GCs, a subset of PMSG- or hCG-induced genes showed a differential expression in Er{beta}null GCs. These latter ER{beta}-regulated genes included previously known FSH or LH target genes including Lhcgr, Cyp11a1, Cyp19a1, Pgr, Runx2, Egfr, Kiss1, and Ptgs2, which are involved in follicle development, oocyte maturation, and ovulation. We also identified novel ER{beta}-regulated genes including Jaml, Galnt6, Znf750, Dusp9, Wnt16, and Mageb16 that failed to respond to gonadotropin stimulation in Er{beta}null GCs. Our findings indicate that the gonadotropin-induced spatiotemporal pattern of gene expression is essential for ovarian follicle maturation beyond the antral stage. However, expression of a subset of those gonadotropin-induced genes is dependent on transcriptional regulation by ER{beta}.

developmental biology↗

bioRxiv2020

SATB homeobox 1 regulated genes in the mouse ectoplacental cone are important for placental development

SATB homeobox 1 (SATB1) is abundantly expressed in the stem-state of trophoblast cells but downregulated during trophoblast differentiation. It is also expressed in high levels in the mouse ectoplacental cones (EPCs). We detected that SATB1 is involved in maintaining the self-renewal of trophoblast stem cells and inhibiting trophoblast differentiation. In this study, we have identified SATB1-regulated genes in the mouse EPC and analyzed their potential functions. A total of 1618 differentially expressed genes were identified in Satb1null EPCs by mRNA sequencing. Remarkably 90% of the differentially expressed genes were found to be upregulated in Satb1null EPCs suggesting a transcriptional repressor role of SATB1 in mouse trophoblast cells. Ingenuity Pathway Analyses demonstrated that the differentially expressed genes in Satb1null EPCs are particularly linked to WNT and TGF{beta} signaling pathways, which regulate self-renewal of stem cells and cell differentiation. Moreover, twenty-six of the EPC genes that are known to be involved in placental development including Eomes, Epas1, Fgfr2, Cdkn1c, and Plac9 were found dysregulated in Satb1null EPCs due to the loss of SATB1 expression. These genes are particularly involved in the formation of labyrinthine zone. Our results emphasize that SATB1-regulated genes in the mouse EPC contribute to key roles in the regulation of trophoblast differentiation and placental development.

developmental biology↗

bioRxiv2020

A long-range chromatin interaction regulates SATB homeobox 1 gene expression in trophoblast stem cells

SATB homeobox proteins are important regulators of developmental gene expression. Among the stem cell lineages determined during early embryonic development, trophoblast stem (TS) cells exhibit robust SATB expression. Both SATB1 and SATB2 act to maintain trophoblast stem-state. However, the molecular mechanisms that regulate TS-specific Satb expression are not yet known. We identified Satb1 variant 2 as the predominant transcript in trophoblasts. Histone marks, and RNA polymerase II occupancy in TS cells indicated active state of the promoter. A novel cis-regulatory region with active histone marks was identified [~]21kbp upstream of variant 2 promoter. CRISPR/Cas9 mediated disruption of this sequence decreased Satb1 expression in TS cells and chromatin conformation capture confirmed looping of this regulatory region into the promoter. Scanning position weight matrices across the enhancer predicted two ELF5 binding sites in close vicinity of SATB1 sites, which were confirmed by chromatin immunoprecipitation. Knockdown of ELF5 downregulated Satb1 expression in TS cells and overexpression of ELF5 increased the enhancer-reporter activity. Interestingly, ELF5 interacts with SATB1 in TS cells, and the enhancer activity was upregulated following SATB overexpression. Our findings indicate that trophoblast-specific Satb1 expression is regulated by long-range chromatin looping of an enhancer that interacts with ELF5 and SATB proteins.

developmental biology↗

bioRxiv2020

ERβ regulated ovarian kisspeptin plays an important role in oocyte maturation

Kisspeptin (KISS1) signaling in the hypothalamic-pituitary (H-P) axis plays essential role in regulating gonadotropin secretion. KISS1 and KISS1 receptor (KISS1R) are also expressed in the ovary; however, the role of intraovarian KISS1 signaling remains largely unclear. Granulosa cell (GC)-specific expression of KISS1, and oocyte-specific expression of KISS1R indicate that GC-derived KISS1 may act on oocytes. Expression of KISS1 in GCs is induced by gonadotropins but it is absent in estrogen receptor {beta} knockout (Er{beta}null) rats. We also observed that gonadotropin stimulation failed to induce maturation of Er{beta}null oocytes. Interestingly, KISS1 treatment of cumulus oocyte complexes (COCs) isolated from antral follicles promotes in vitro maturation of oocytes. Treatment of oocytes with KISS1 induced intracellular Ca2+ release, and increased activation of MAP kinase ERK1/2. KISS1 treatment also induced the expression of oocyte genes that are crucial for differentiation of GCs, and maturation of oocytes. Our findings suggest that ovarian KISS1-signaling plays an important role in gonadotropin induced follicle development and oocyte maturation.

developmental biology↗

bioRxiv2020

A gatekeeping role of ESR2 to maintain the primordial follicle reserve

Over the entire reproductive lifespan in mammals, a fixed number of primordial follicles serve as the source of mature oocytes. Uncontrolled and excessive activation of primordial follicles can lead to depletion of the ovarian reserve. We observed that disruption of ESR2-signaling results in increased activation of primordial follicles in Esr2-null (Esr2-/-) rats. However, follicle assembly was unaffected, and the total number of follicles remained comparable between neonatal wildtype and Esr2-/- ovaries. While the activated follicle counts were increased in Esr2-/- ovary, the number of primordial follicles were markedly decreased. Excessive recruitment of primordial follicles led to premature ovarian senescence in Esr2-/- rats and was associated with reduced levels of serum AMH and estradiol. Disruption of ESR2-signaling through administration of a selective antagonist (PHTPP) increased the number of activated follicles in wildtype rats, whereas a selective agonist (DPN) decreased follicle activation. In contrast, primordial follicle activation was not increased in the absence of ESR1 indicating that the regulation of primordial follicle activation is ESR2-specific. Follicle activation was also increased in Esr2-mutants lacking the DNA-binding domain, suggesting a role for the canonical transcriptional activation function. Both primordial and activated follicles express ESR2 suggesting a direct regulatory role for ESR2 within these follicles. We also detected that loss of ESR2 augmented the activation of AKT, ERK and mTOR pathways. Our results indicate that the lack of ESR2 upregulated both granulosa and oocyte factors, which can facilitate AKT and mTOR activation in Esr2-/- ovaries leading to increased activation of primordial follicles.

developmental biology↗