NLRP3 Inflammasome Activation of Mast Cells by Estrogen via the Nuclear-Initiated Signaling Pathway Contributes to the Development of Endometriosis

Endometriosis is an estrogen-dependent gynecological disease. The pathogenesis of endometriosis remains controversial, although it is generally accepted that the inflammatory immune response plays a crucial role in this process. Mast cells (MCs) are multifunctional innate immune cells that accumulate in endometriotic lesions. However, the molecular mechanism by which estrogen modulates MCs in the development of endometriosis is not well understood. Here we report that estrogen can induce the expression of NOD-like receptor family pyrin domain containing 3 (NLRP3) through estrogen receptor (ER)-α via the estrogen responsive element (ERE) in MCs. Such transcriptional regulation is necessary for the activation of NLRP3 inflammasome and the production of mature interleukin (IL)-1β in MCs. Targeted inhibition of NLRP3 significantly restrained lesion progression and fibrogenesis in a mouse model of endometriosis. Collectively, these findings suggest that MCs contribute to the development of endometriosis through NLRP3 inflammasome activation mediated by nuclear-initiated estrogen signaling pathway.

Estrogen Receptor β Activation Inhibits Colitis by Promoting NLRP6-Mediated Autophagy

Estrogen receptor β (ERβ) and NLRP6 are highly expressed in intestinal tissues. Loss of ERβ and NLRP6 exacerbate colitis in mouse models. However, the underlying mechanisms are incompletely understood. Here, we report that ERβ attenuates inflammation by inducing NLRP6-mediated autophagy. Specifically, ERβ directly activates the NLRP6 gene expression via binding to estrogen responsive element (ERE) of Nlrp6 gene promoter. ERβ also physically interacts with the NLRP6 nucleotide-binding domain and promotes NLRP6 inflammasome assembly. The ERβ-NLRP6 axis then interacts with multiple autophagy-related proteins including ULK1, BECN1, ATG16L1, LC3B, p62 to affect the autophagosome biogenesis and control autophagic flux. Finally, NLRP6-mediated autophagy suppresses the inflammatory response by promoting the K48-linked polyubiquitination of ASC, Casp-1 p20, IL-1β, TNF-α, and prohibitin-2. Thus, ERβ-NLRP6 direct an anti-inflammatory response by promoting autophagy. Our work uncovers an ERβ-NLRP6-autophagy pathway as an unrecognized regulatory mechanism that maintains intestinal epithelial cell homeostasis and facilitates tissue repair in colitis.

Real-Time Challenging of ERα Y537S Mutant Transcriptional Activity in Living Cells

Metastatic estrogen receptor α (ERα)-expressing breast cancer (BC) occurs after prolonged patient treatment with endocrine therapy (ET) (e.g., aromatase inhibitors—AI; 4OH-tamoxifen—4OH-Tam). Often these metastatic BCs express a mutated ERα variant (e.g., Y537S), which is transcriptionally hyperactive, sustains uncontrolled proliferation, and renders tumor cells insensitive to ET drugs. Therefore, new molecules blocking hyperactive Y537S ERα mutation transcriptional activity are requested. Here we generated an MCF-7 cell line expressing the Y537S ERα mutation stably expressing an estrogen-responsive element (ERE) promoter, which activity can be monitored in living cells. Characterization of this cell line shows both hyperactive basal transcriptional activity with respect to normal MCF-7 cells, which stably express the same ERE-based promoter and a decreased effect of selective ER downregulators (SERDs) in reducing Y537S ERα mutant transcriptional activity with respect to wild type ERα transcriptional activity. Kinetic profiles of Y537S ERα mutant-based transcription produced by both drugs inducing receptor degradation and siRNA-mediated depletion of specific proteins (e.g., FOXA1 and caveolin1) reveals biphasic dynamics of the inhibition of the receptor-regulated transcriptional effects. Overall, we report a new model where to study the behavior of the Y537S ERα mutant that can be used for the identification of new targets and pathways regulating the Y537S ERα transcriptional activity.

Estrogen Induces Selective Transcription of Caveolin1 Variants in Human Breast Cancer through Estrogen Responsive Element-Dependent Mechanisms

The estrogen receptor (ER) signaling regulates numerous physiological processes mainly through activation of gene transcription (genomic pathways). Caveolin1 (CAV1) is a membrane-resident protein that behaves as platform to enable different signaling molecules and receptors for membrane-initiated pathways. CAV1 directly interacts with ERs and allows their localization on membrane with consequent activation of ER-non-genomic pathways. Loss of CAV1 function is a common feature of different types of cancers, including breast cancer. Two protein isoforms, CAV1α and CAV1β, derived from two alternative translation initiation sites, are commonly described for this gene. However, the exact transcriptional regulation underlying CAV1 expression pattern is poorly elucidated. In this study, we dissect the molecular mechanism involved in selective expression of CAV1β isoform, induced by estrogens and downregulated in breast cancer. Luciferase assays and Chromatin immunoprecipitation demonstrate that transcriptional activation is triggered by estrogen-responsive elements embedded in CAV1 intragenic regions and DNA-binding of estrogen-ER complexes. This regulatory control is dynamically established by local chromatin changes, as proved by the occurrence of histone H3 methylation/demethylation events and association of modifier proteins as well as modification of H3 acetylation status. Thus, we demonstrate for the first time, an estrogen-ERs-dependent regulatory circuit sustaining selective CAV1β expression.

Efp as a primary estrogen‐responsive gene in human breast cancer.

We have previously isolated the efp (estrogenresponsivefinger protein) that is required for the normalestrogen-induced cell proliferation. Here, we show the genomicorganization of the human efp gene which consists of nine exons.The efp mRNA was expressed in human breast tumors and theestrogen-induced expression of the efp was found in MCF-7human breast cancer cells. Moreover, efp promoter activity wasenhanced through the estrogen-responsive element dependent onestrogen and estrogen receptor. These results suggest that the efpcan mediate estrogen actions such as cell growth in human breastcancer as a primary responsive gene.

17β-estradiol binding to ERα promotes the progression of prolactinoma through estrogen-response element-induced CaBP-9k up-regulation

17β-estradiol (E2) is considered to be an important instigator of prolactinoma, and can positively regulate the expression of calbindin-D9k (CaBP-9k) which contains an estrogen responsive element (ERE) via estrogen receptors (ERs). However, the detailed mechanism of E2 in promoting CaBP-9k expression and their roles in prolactinoma progression remain unclear. Here, we aimed to characterize it. The luciferase gene reporter assay with luc-ERE transfection showed that E2 treatment significantly enhanced the transcriptional level of CaBP-9k, whereas CaBP-9k activity was reduced when GH3 and MMQ cells were treated with AZD9496, an antagonist of ERα. E2 treatment increased the protein expressions of CaBP-9k and ERα but not ERβ, whereas this effect was also abolished when cells were treated with AZD9496. Besides, immunoprecipitation (IP) and immunofluorescence assays demonstrated that CaBP-9k could directly interact with ERα not ERβ, and Chromatin IP (ChIP) assay showed that ERα could bind to ERE of the CaBP-9k promoter. Moreover, cell counting kit-8 (CCK-8) and flow cytometry assays showed that E2 treatment significantly enhanced cell viability and inhibited cell apoptosis, but these effects were all abolished when ERα was down-regulated by short hairpin RNA (shRNA) or inhibited by AZD9496, as well as CaBP-9K suppression in both GH3 and MMQ cell lines. Taken together, these findings indicated that E2 stimulation promoted prolactin cell proliferation and inhibited cell apoptosis through ERα-induced CaBP-9k up-regulation, which then accelerated the advanced progression of prolactinoma.

Neuropeptide B mediates female sexual receptivity in medaka fish, acting in a female-specific but reversible manner

Male and female animals display innate sex-specific mating behaviors. In teleost fish, altering the adult sex steroid milieu can effectively reverse sex-typical mating behaviors, suggesting remarkable sexual lability of their brains as adults. In the teleost medaka, neuropeptide B (NPB) is expressed female-specifically in the brain nuclei implicated in mating behavior. Here, we demonstrate that NPB is a direct mediator of estrogen action on female mating behavior, acting in a female-specific but reversible manner. Analysis of regulatory mechanisms revealed that the female-specific expression of NPB is dependent on direct transcriptional activation by estrogen via an estrogen-responsive element and is reversed in response to changes in the adult sex steroid milieu. Behavioral studies of NPB knockouts revealed that female-specific NBP mediates female receptivity to male courtship. The female-specific NPB signaling identified herein is presumably a critical element of the neural circuitry underlying sexual dimorphism and lability of mating behaviors in teleosts.

Neuropeptide B mediates female sexual receptivity in medaka fish, acting in a female-specific but reversible manner

Male and female animals display innate sex-specific mating behaviors. Among vertebrates, teleosts are unique in that altering the adult sex steroid milieu can reverse sex-typical mating behaviors, suggesting sexual lability of their brains. In the teleost medaka, neuropeptide B (NPB) is expressed female-specifically in the brain nuclei implicated in mating behavior. Here, we demonstrate that NPB is a direct mediator of estrogen action on female mating behavior, acting in a female-specific but reversible manner. Analysis of regulatory mechanisms revealed that the female-specific expression of NPB is dependent on direct transcriptional activation by estrogen via an estrogen-responsive element and is reversed in response to changes in the adult sex steroid milieu. Behavioral studies of NPB knockouts revealed that female-specific NBP mediates female receptivity to male courtship. The female-specific NPB signaling identified herein is presumably a critical element of the neural circuitry underlying sexual dimorphism and lability of mating behaviors in teleosts.