Calcium and Cadmium Activate ESRRB to Mediate Cell Stemness and Pluripotency

Estrogen-related receptor beta (ESRRB) is thought to be an orphan receptor that functions as a transcription factor, pioneer factor, and mitotic bookmarker to regulate cell stemness, pluripotency, and differentiation. This study 1) investigates whether calcium and cadmium activation of ESRRB regulates signaling pathways of stemness and pluripotency, 2) explores the transcriptomic and biological alterations of metal activation of ESRRB, and 3) reveals the underlying mechanisms by which metals activate ESRRB. In HEK293T cells, treatment with calcium and cadmium increased the expression of ESRRB-regulated genes that was blocked by an ESRRB antagonist. In the breast cancer cell line MDA-MB-453, treatment with calcium, cadmium, or a synthetic agonist also increased expression of ESRRB-regulated genes that was blocked by the antagonist, enhanced ESRRB nuclear localization, increased the recruitment of RNA polymerase 2 to estrogen-related receptor response elements, enhanced cell stemness and proliferation pathways, and induced the expression of estrogen receptor alpha. Mutational analysis and molecular docking identified potential metal interaction sites within ESRRB's ligand-binding domain. Together, these results suggest calcium acts as a natural ligand for ESRRB and cadmium, which mimics calcium, activate ESRRB to mediate cell stemness and pluripotency. Overall design: RNA-seq profiling of MDA-MB-453 cells with treatment of calcium, cadmium, DES (ESRRB antagonist), DES+calcium, DES+cadmium, or GSK4716 (ESRRB agonist) for 24 hours, plus no-treatment controls.