Deciphering the molecular mechanisms of STIM1 and Orai1: coronavirus cleaves STIM1 to evade innate immune and Orai1 negatively regulates KLF1 in terminal erythropoiesis

Abstract

This dissertation consists of two independent studies centered on major component of SOCE, STIM1 and Orai1, which mediate distinct regulatory mechanisms in host–virus interaction and erythropoiesis. In the first study, I investigated how human coronaviruses exploit host cellular machinery to evade antiviral immunity. I identified STIM1, a key Ca²⁺ sensor in the endoplasmic reticulum, as a previously unrecognized substrate of the coronavirus 3CL protease. Cleavage of STIM1 at residue Q496 generates two fragments that acquire novel proviral functions. The STIM1 NT inhibits MAVS aggregation and MAVS–TRAF2–TBK1 signalosome formation, whereas the STIM1 CT interferes with IKKα-mediated p65 phosphorylation through its interaction with HSP70. Together, these mechanisms cooperatively suppress interferon-β production and attenuate host antiviral responses, revealing a new strategy by which coronaviruses subvert host immunity through targeted cleavage of STIM1. The second study focuses on the role of Orai1-mediated Ca²⁺ signaling in terminal erythroid maturation. Erythropoietin (EPO) and the master transcription factor Krüppel-like factor 1 (KLF1) orchestrate the progression of erythropoiesis; however, the contribution of Ca²⁺ signaling to this process has remained poorly defined. I found Orai1 as a novel EPO-responsive Ca²⁺ channel that functions as a dynamic regulatory toggle controlling KLF1 transcription. In early erythroid differentiation, EPO- induced Orai1 activity suppresses KLF1 expression via Ca²⁺-dependent NFAT2 activation, transiently pausing maturation. As differentiation proceeds, Orai1 expression declines, relieving NFAT2-mediated repression and allowing EPO–STAT5-dependent activation of KLF1 to promote terminal erythropoiesis. Functional disruption of Orai1, by R91W mutation or CRISPR/Cas9-mediated knockout, enhances KLF1 expression, globin synthesis, and enucleation efficiency in HUDEP-2, UCB-, and hPSC-derived erythroid cells. Collectively, this dissertation delineates two distinct molecular strategies—one utilized by viruses to undermine host immune defenses and another employed by developing erythroblasts to coordinate maturation—underscoring the versatility of calcium-related regulatory pathways in controlling cellular function and disease processes.