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Abstract
TNFα contributes to type 1 diabetes pathogenesis by impairing beta cell function and driving beta cell apoptosis. How TNFα directs these processes is not fully understood. Of note, baculoviral-IAP-repeat-containing (BIRC) proteins associate with the TNF complex to control down stream signaling but their function in beta cells is not known. Following TNF receptor ligation, Birc3 but not Birc1,2,4,5 or -6 was induced in mouse islets and MIN6 cells. Contact with TNFα triggers signaling cascades converging on the NF-κB, JNK, and p38 pathways that regulate inflammatory genes. The Birc3-promoter harboured three NF-κB and an AP-1 binding sites. Induction of endogenous Birc3 by TNFα was completely blocked by NF-κB inhibitors - thus NF-κB regulates Birc3 transcription. Further, BIRC3 gain-of-function induced activation of an NF-κB reporter and potentiated TNFα-induced NF-κB activation. Surprisingly, Birc3-/- islets showed delayed but functional NF-κB activation and exhibited dysregulated TNFα-induced Ccl2, Cxcl10 and Icam-1. Some data demonstrate BIRC2 can compensate for loss of BIRC3. Expectedly, Birc2-/-Birc3-/- islets showed markedly delayed IκBα degradation kinetics; suggesting that BIRC2/3 are crucial for NF-κB signaling. Remarkably, Birc2-/- and Birc3-/- islets showed unexpected phenotypes independent of NF-κB. Genetic analysis showed Birc2-/- islets exhibited increased basal Ccl2 and Cxcl10 expression. Cell signaling was further dysregulated by the addition of BIRC3 deficiency. Birc2-/-Birc3-/- islets exhibited increased basal A20, Icam-1, Ccl2 and Cxcl10, which were blocked by antagonizing JNK but not NF-κB or p38 pathway. Moreover, Birc2-/-Birc3-/- islets showed hyperphosphorylation of the JNK-target, c-Jun. BIRC proteins ubiquitinate substrates targeting them for proteosomal degradation. Proteosome inhibition in MIN6 cells mimicked the Birc2-/-Birc3-/- islet phenotype – increased basal Cxcl10, Ccl2 and Icam-1 expression with c-Jun-hyperphosphorylation. Birc2-/-Birc3-/- islets transplanted into diabetic-allogeneic recipients showed rapid loss of function indicative of dysregulated stress response. We propose that loss of BIRC2/3 uncoupled molecular control of inflammatory genes from TNF-signaling cascades. These present data demonstrate a novel, cell specific role for BIRC2 and BIRC3 as molecular rheostat that fine-tune NF-κB and JNK signaling to ensure transcriptional responses are appropriately matched to extra-cellular inputs. These pathways may be critical for beta cell stress responses.