Communication between the endoplasmic reticulum (ER) and mitochondrion is important for bioenergetics and cellular survival. The ER supplies Ca²⁺ directly to mitochondria via inositol 1,4,5-trisphosphate receptors (IP3Rs) at close contacts between the two organelles referred to as mitochondrion-associated ER membrane (MAM). We found here that the ER protein sigma-1 receptor (Sig-1R), which is implicated in neuroprotection, carcinogenesis, and neuroplasticity, is a Ca²⁺-sensitive and ligand-operated receptor chaperone at MAM. Normally, Sig-1Rs form a complex at MAM with another chaperone, BiP. Upon ER Ca²⁺ depletion or via ligand stimulation, Sig-1Rs dissociate from BiP, leading to a prolonged Ca²⁺ signaling into mitochondria via IP3Rs. Sig-1Rs can translocate under chronic ER stress. Increasing Sig-1Rs in cells counteracts ER stress response, whereas decreasing them enhances apoptosis. These results reveal that the orchestrated ER chaperone machinery at MAM, by sensing ER Ca²⁺ concentrations, regulates ER-mitochondrial interorganellar Ca²⁺ signaling and cell survival.