Homocysteine-inducible, endoplasmic reticulum (ER) stress-inducible, ubiquitin-like domain member 1 (HERPUD1), an ER resident protein, is upregulated in response to ER stress and Ca²⁺ homeostasis deregulation. HERPUD1 exerts cytoprotective effects in various models, but its role during oxidative insult remains unknown. The aim of this study was to investigate whether HERPUD1 contributes to cytoprotection in response to redox stress and participates in mediating the stress-dependent signaling pathways. Our data showed that HERPUD1 protein levels increased in HeLa cells treated for 30 min with H₂O₂ or angiotensin II and in aortic tissue isolated from mice treated with angiotensin II for 3 weeks. Cell death was higher in HERPUD1 knockdown (sh-HERPUD1) in HeLa cells treated with H₂O₂ in comparison with control (sh-Luc) HeLa cells. This effect was abolished by the intracellular Ca²⁺ chelating agent BAPTA-AM or the inositol 1,4,5-trisphosphate receptor (ITPR) antagonist xestospongin B, suggesting that the response to H₂O₂ was dependent on intracellular Ca²⁺ stores and the ITPR. Ca²⁺ kinetics showed that sh-HERPUD1 HeLa cells exhibited greater and more sustained cytosolic and mitochondrial Ca²⁺ increases than sh-Luc HeLa cells. This higher sensitivity of sh-HERPUD1 HeLa cells to H₂O₂ was prevented with the mitochondrial permeability transition pore inhibitor cyclosporine A. We concluded that the HERPUD1-mediated cytoprotective effect against oxidative stress depends on the ITPR and Ca²⁺ transfer from the ER to mitochondria.