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The liver helps maintain energy homeostasis by synthesizing and storing glucose and lipids. Gonadal steroids, particularly estrogens, play an important role in regulating metabolism. As estrogens are considered female hormones, metabolic disorders related to the disruption of estrogen signaling have mostly been studied in females. Estrogen receptor alpha (ESR1) is the predominant receptor in both the male and female liver, and it mediates the hepatic response to estrogens. Loss of ESR1 increases weight gain and obesity in female rats, while reducing the normal growth in males. Although Esr1-/- male rats have a reduced body weight, they exhibit increased adipose deposition and impaired glucose tolerance. We further investigated whether these metabolic disorders in Esr1-/- male rats were linked with the loss of transcriptional regulation by ESR1 in the liver. To identify the ESR-regulated genes, RNA-sequencing was performed on liver mRNAs from wildtype and Esr1-/- male rats. Based on an absolute fold change of ≥2 with a p-value ≤ 0.05, a total of 706 differentially expressed genes were identified in the Esr1-/- male liver: 478 downregulated, and 228 upregulated. Pathway analyses demonstrate that the differentially expressed genes include transcriptional regulators (Cry1, Nr1d1, Nr0b2), transporters (Slc1a2), and regulators of biosynthesis (Cyp7b1, Cyp8b1), and hormone metabolism (Hsd17b2, Sult1e1). Many of these genes are also integral parts of the lipid and carbohydrate metabolism pathways in the liver. Interestingly, certain critical regulators of the metabolic pathways displayed a sexual dimorphism in expression, which may explain the divergent weight gain in Esr1-/- male and female rats despite common metabolic dysfunctions., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Estrogen signaling plays an important role in the pathophysiology of prostatic hyperplasia. While signaling through estrogen receptor alpha (ESR1) increases proliferation of stromal cells, estrogen receptor beta (ESR2) plays an anti-proliferative and differentiating role in glandular epithelium. Disruption of ESR2 signaling resulted in prostatic glandular hyperplasia in the rat. To identify the ESR2-target genes, and the molecular mechanisms involved, we performed RNA-seq analyses in prostate glands of Esr2 knockout ( Esr2 -/-) and age matched wildtype rats. The raw data were analyzed using CLC genomics workbench. High quality RNA-seq reads were aligned to the Rattus norvegicus genome. Differentially expressed genes were identified based on an absolute fold change of 2 with p Value ≤0.05. Of the total 32,623 genes detected, 824 were differentially expressed in Esr2 -/- prostate glands, 550 downregulated and 274 upregulated. Pathway analyses identified altered expression of genes involved in epithelial proliferation and benign tumor formation.

Estrogens are traditionally considered to be female sex steroid hormones and most of the studies examining estrogen regulation of metabolic function in the liver have been conducted in females. However, the liver expresses high levels of estrogen receptor alpha (ESR1) in both males and females, which mediates the hepatic response to estrogens. In this data article, we investigated whether metabolic disorders in Esr1 knockout ( Esr1-/- ) male rats were linked with loss of transcriptional regulation by ESR1 in liver. To identify the ESR1 regulated genes in the mutant liver, RNA-sequencing was performed on liver RNAs purified from young male rats. The raw data were analyzed using the CLC Genomics Workbench and high-quality RNA-sequencing reads were aligned to the Rattus norvegicus genome. Transcriptome data obtained from Esr1-/- liver RNAs were compared to that of wild type rats. Based on an absolute fold change of 2 with a p -value ≤ 0.05, a total of 618 differentially expressed genes were identified in the Esr1-/- male liver. Pathway analyses demonstrated that the majority of differentially expressed genes are regulators of carbohydrate and lipid metabolism in the liver. These differentially expressed genes and their potential roles were further examined in a companion manuscript, "Disruption of ESR1 alters the expression of genes regulating hepatic lipid and carbohydrate metabolism in male rats" (Khristi et al., 2018).

Hypothalamic expression of Kiss1 plays an essential role in the onset of puberty, gonadal development, and ovulation. Estrogens regulate the expression of Kiss1 in the hypothalamus through estrogen receptor-α. Kiss1 is also expressed in the ovary, where its expression correlates with the onset of puberty and progression of the estrous cycle. To date, estrogen regulation of Kiss1 expression in the ovary has not been investigated. We recently observed that gonadotropin-induced Kiss1 expression was absent in Esr2-null rat ovaries even though Esr1 was present. Wild-type granulosa cells abundantly expressed Kiss1 and oocytes expressed the Kiss1 receptor. We characterized estrogen receptor-β (ESR2) regulation of Kiss1 expression in granulosa cells by identifying granulosa cell-specific transcript variants and potential regulatory regions. The Kiss1 promoter, an upstream enhancer, and a downstream enhancer all possessed conserved estrogen response elements (EREs) and showed active histone marks in gonadotropin-stimulated granulosa cells. The transcriptionally active Kiss1 promoter, as well as the enhancers, also revealed enrichment for ESR2 binding. Furthermore, activity of a Kiss1 promoter construct was induced after overexpression of ESR2 and was blocked upon mutation of an ERE within the promoter. Finally, pregnant mare serum gonadotropin and human chorionic gonadotropin administration induced phosphorylation of ESR2 and upregulated the AP-1 proteins FOSL2 and JUNB in granulosa cells. Activated MAPK ERK2 was associated with the ESR2 phosphorylation in granulosa cells, and AP-1 factors could synergistically activate the Kiss1 promoter activity. These gonadotropin-induced changes paralleled Kiss1 expression in granulosa cells. We conclude that gonadotropin-stimulated Kiss1 expression in granulosa cells is dependent on both the activation of ESR2 and the upregulation of AP-1.

Estrogen receptor 2 (ESR2) plays a critical role in folliculogenesis and ovulation. Disruption of ESR2-function in the rats results in female infertility due to failure of ovulation. Ovulation failure occurred in two distinct rat models, a null mutant and a DNA binding domain (DBD) mutant of ESR2, indicating that transcriptional regulation by ESR2 is indispensable for ovulation. To define the regulatory role of ESR2 in preovulatory follicular maturation and ovulation, we investigated ovarian responsiveness to exogenous gonadotropins in prepubertal females. Granulosa cells (GCs) play a vital role in follicle maturation and ovulation, and ESR2-dependent estrogen signaling is predominant in GCs, therefore, we examined the differential expression of gonadotropin-induced genes in GCs. Of 32,623 genes detected by RNA-sequencing, 1696 were differentially expressed in Esr2-mutant rats (789 downregulated, and 907 upregulated, absolute fold change 2, FDR p < 0.05). Molecular pathway analyses indicated that these differentially expressed genes are involved in steroidogenesis, follicle maturation, and ovulation. Many of these genes are known regulators of ovarian function and a subset were also disrupted in Esr2-mutant mice. Interestingly, Kiss1 was identified as one of the differentially expressed genes implicating a potential role within the follicle and its regulation by ESR2. Our findings indicate that ESR2 regulates key genes in GCs that are essential for follicle maturation and ovulation in the rat., (Copyright © 2018 Elsevier B.V. All rights reserved.)

RNA seq analyses were performed in granulosa cells (GCs) collected from gonadotropin treated ESR2 mutant rats. Data obtained from a null mutant with Esr2 exon 3 deletion (∆3) and another DNA binding domain (DBD) mutant with exon 4 deletion (∆4) were compared to that of wildtype (WT) rats. The raw data were analyzed using CLC genomics workbench. High quality RNA-sequencing reads were aligned to the Rattus norvegicus genome. Differentially expressed genes in ∆3 or ∆4 Esr2 -mutant GCs were identified based on the following criteria: FDR p-Value ≤0.05 and an absolute fold change of 2. Fewer differentially expressed genes were identified in ∆3 compared to the ∆4 mutant group. As both mutant groups demonstrated a common phenotype of ovulation failure, differentially expressed genes common to both in ∆3 and ∆4 mutant rats were emphasized and further analyzed in the companion article "ESR2 regulates granulosa cell genes essential for follicle maturation and ovulation" [1].