Chronic inflammation and pro-inflammatory cytokines are important mediators of pancreatic β-cell destruction in type 1 diabetes (T1D). We presently show that the cytokines IL-1β+ IFN-γ and different ER stressors activate the Bcl-2 homology 3 (BH3)-only member death protein 5 (DP5)/harakiri (Hrk) resulting in β-cell apoptosis. Chemical ER stress-induced DP5 upregulation is JNK/c-Jun-dependent. DP5 activation by cytokines also involves JNK/c-Jun phosphorylation and is antagonized by JunB. Interestingly, cytokine-inducted DP5 expression precedes ER stress: mitochondrial release of cytochrome c and ER stress are actually a consequence of enhanced DP5 activation by cytokine-mediated nitric oxide formation. Our findings show that DP5 is central for β-cell apoptosis after different stimuli, and that it can act up-and downstream of ER stress. These observations contribute to solve two important questions, namely the mechanism by which IL-1β+ IFN-γ induce β-cell death and the nature of the downstream signals by which ER stress ‘convinces’ β-cells to trigger apoptosis.