Prostate cancer (PCa) is the second leading cause of mortality and a leading cause of malignant tumors in males. Prostate cancer stem cells (PCSCs) are likely the responsible cell types for cancer initiation, clinical treatment failure, tumor relapse, and metastasis. Estrogen receptor alpha (ERα) is mainly expressed in the basal layer cells of the normal prostate gland and has key roles in coordinating stem cells to control prostate organ development. Here, we investigated the roles of the estrogen-ERα signaling pathway in regulating PCSCs. Correlation of CD49f and ERα/NOTCH1 was analyzed in human clinical datasets and tissue samples. Flow cytometry was used to sort CD49fHi and CD49fLow cells. EZH2 recruitment by ERα and facilitation of ERα binding to the NOTCH1 promoter was validated by Co-IP and ChIP. Primary tumor growth, tumor metastasis and sensitivity to 17β-estradiol (E2) inhibitor (tamoxifen) were evaluated in castrated mice. ERα expression was significantly higher in CD49fHi prostate cancer basal stem-like cells (PCBSLCs), which showed basal and EMT features with susceptibility to E2 treatment. ERα-induced estrogen effects were suggested to drive the NOTCH1 signaling pathway activity via binding to the NOTCH1 promoter. Moreover, EZH2 was recruited by ERα and acted as a cofactor to assist ERα-induced estrogen effects in regulating NOTCH1 in PCa. In vivo, E2 promoted tumor formation and metastasis, which were inhibited by tamoxifen. Our results implicated CD49f+/ERα + prostate cancer cells associated with basal stem-like and EMT features, named EMT-PCBSLCs, in heightened potential for promoting metastasis. NOTCH1 was regulated by E2 in CD49fHi EMT-PCBSLCs. These results contribute to insights into the metastatic mechanisms of EMT-PCBSLCs in PCa.

Background: Resistance to androgen deprivation therapy remains a major challenge for the clinical treatment of patients with castration-resistant prostate cancer (CRPC). CYP1B1, a critical enzyme that catalyzes the conversion of estradiol to 4-Hydroxy-17β-estradiol (4-OHE2), has been reported to promote the development and progression of hormone-related cancer, but its role in CRPC is unclear. Methods: To explore the underlying mechanism which CYP1B1 promotes the prostate cancer stem cells (PCSCs) characteristics, bioinformatics analyses of human clinical prostate cancer (PCa) datasets were performed. CYP1B1, IL6, and estrogen receptor-α (ERα) expression levels were evaluated in PCa and CRPC tissues via immunohistochemistry. The high-performance liquid chromatography-mass spectrometry assay was carried out to examine intracellular 4-OHE2 levels. Serum-free suspension culture and flow cytometry assays were performed to evaluate PCSCs. Chromatin immunoprecipitation was used to validate that 4-OHE2 recruited ERα to the IL6 promoter. Results: CYP1B1 expression was significantly increased in CRPC tissues and androgen-independent PCa cell lines. CYP1B1⁺ PCa cells were significantly enriched in bicalutamide-treated LNCaP cells, and CYP1B1 knockdown reduced the cell viability under bicalutamide treatment. In addition, CYP1B1 knockdown decreased the intracellular 4-OHE2 concentration, accompanied by reduced PCSC characteristics. In PCa cells, 4-OHE2 stimulated ERα transcriptional activity and upregulated the expression of IL6 and downstream genes of the IL6-STAT3 signaling. 4-OHE2 increased cell viability under bicalutamide treatment and promoted PCSC characteristics, while IL6 neutralizing antibody reversed these effects. Mechanistically, siERα and the ER antagonist ICI182780 significantly attenuated 4-OHE2-induced IL6 expression, and 4-OHE2 promoted the binding of ERα to the estrogen response element of the IL6 promoter. Conclusions: Our findings indicate that CYP1B1-catalyzed 4-OHE2 enhanced PCSC characteristics and attenuated bicalutamide sensitivity by ERα-mediated the IL6-STAT3 pathway activation. Our study further emphasizes the role of CYP1B1 in castration resistance and illustrates a novel mechanism of CRPC development. Graphical

Embryonic stem cells (ESCs) favor glycolysis over oxidative phosphorylation for energy production, and glycolytic metabolism is critical for pluripotency establishment, maintenance, and exit. However, an understanding of how glycolysis regulates the self-renewal and differentiation of ESCs remains elusive. Here, we demonstrated that protein lactylation, regulated by intracellular lactate, contributes to the self-renewal of ESCs. We further showed that Esrrb, an orphan nuclear receptor involved in pluripotency maintenance and extraembryonic endoderm stem cell (XEN) differentiation, is lactylated on K228 and K232. The lactylation of Esrrb enhances its activity in promoting ESC self-renewal in the absence of the LIF and XEN differentiation of ESCs by increasing its binding at target genes. Our studies reveal the importance of protein lactylation in the self-renewal and XEN differentiation of ESCs, and the underlying mechanism of glycolytic metabolism regulating cell fate choice.