<p>Sepsis is associated with a pronounced but poorly understood endoplasmic reticulum stress (ERS) response. In this study, we found that death-associated protein kinase 2 (DAPK2), a calcium/calmodulin-regulated serine/threonine kinase, exhibits elevated expression in macrophages from patients with sepsis and from septic mice. Macrophage DAPK2 expression is transcriptionally upregulated through the activation of the Toll-like receptor 4 (TLR4)–myeloid differentiation primary response 88 (MyD88)–nuclear factor-κB (NF-κB) pathway. Macrophage-specific deletion of DAPK2 attenuated sepsis severity and mitigated the ERS response. Using liquid chromatography–tandem mass spectrometry (LC–MS/MS), we identified heat shock protein family A member 5 (HSPA5) as a binding partner for DAPK2. Because DAPK2 function had been previously associated with the kinase activity, we speculated that it might control ERS of macrophages through HSPA5 phosphorylation. Further investigation indeed revealed that DAPK2 phosphorylates HSPA5 at serine-588, which promotes the proteasomal degradation of HSPA5 and subsequently leads to the activation of inositol-requiring enzyme 1α (IRE1α). Inhibition of HSPA5 exacerbated sepsis in mice with macrophage-specific DAPK2 deficiency; however, this effect was abrogated by the deactivation of IRE1α. In conclusion, our findings demonstrate that DAPK2 propagates macrophage ERS through the HSPA5–IRE1α axis during systemic infection, suggesting this pathway as a potential therapeutic target in sepsis.</p>