Nuclear factor-kappaB translocation mediates double-stranded ribonucleic acid-induced NIT-1 beta-cell apoptosis and up-regulates caspase-12 and tumor necrosis factor receptor-associated ligand (TRAIL).
The mechanism of induction of apoptosis by double-stranded RNA (dsRNA) is not fully characterized. The dsRNA is normally present in extremely low quantities in cells, but following infection with RNA viruses, large quantities of the dsRNA viral replicative intermediate may be produced triggering the antiviral response as well as cell death. In this report, transfection of polyinosinic-polycytidylic acid [poly(I:C)] into NIT 1 cells has been used as a model of intracellular dsRNA-induced beta-cell apoptosis. At 18 h post transfection, 45% of the cells were apoptotic as indicated by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end-labeling (TUNEL) staining, and this was accompanied by an increase in nuclear factor kappaB (NF-kappaB) p50/p65 nuclear translocation and cleavage of caspases 3 and 8. The NF-kappaB inhibitor peptide, SN50, significantly reduced caspase-3 activity and the percentage of TUNEL-positive cells, substantiating a role for NF-kappaB in inducing intracellular dsRNA-mediated apoptosis. Concomitantly, RNA-dependent protein kinase activity was observed at 3 h post transfection along with phosphorylation and degradation of inhibitory kappaB-alpha. Expression of TRAIL (TNF-related apoptosis-inducing ligand), Fas, IL-15, and caspase-12 mRNAs was up-regulated in the presence of poly(I:C) but not when SN50 was also added. In contrast, there was no change detected in Fas, Fas-associated death domain, Bcl-2, Bcl-xl, Bax, p53, or XIAP(X-linked inhibitor of apoptosis protein) expression up to 12 h after poly(I:C) transfection. In addition, caspase-12 was cleaved, and phosphorylation of eukaryotic initiation factor 2alpha occurred, suggesting that an endoplasmic reticulum stress pathway was involved in addition to NF-kappaB induction of an extrinsic pathway, possibly mediated by TNF-related apoptosis-inducing ligand.
Robbins
Marjorie A
MA
Department of Pathology, University of British Columbia, Vancouver, Canada V5Z 4H4.
Maksumova
Lola
L
Pocock
Emma
E
Chantler
Janet K
JK
D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D051017:Apoptosis Regulatory Proteins; D001692:Biological Transport / Q000502:physiology; D053454:Caspase 12; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D061945:Caspase Inhibitors; D020169:Caspases / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015852:Eukaryotic Initiation Factor-2 / Q000378:metabolism; D020672:I-kappa B Proteins / Q000378:metabolism; D019409:Interleukin-15 / Q000235:genetics; D007515:Islets of Langerhans / Q000187:drug effects / Q000502:physiology*; D008055:Lipids / Q000494:pharmacology; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D000072000:NF-KappaB Inhibitor alpha; D016328:NF-kappa B / Q000378:metabolism*; D009569:Nitric Oxide / Q000096:biosynthesis; D011070:Poly I-C / Q000493:pharmacokinetics / Q000494:pharmacology*; D012330:RNA, Double-Stranded / Q000494:pharmacology; D053221:TNF-Related Apoptosis-Inducing Ligand; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000378:metabolism*; D015854:Up-Regulation; D019892:eIF-2 Kinase / Q000378:metabolism; D019014:fas Receptor / Q000235:genetics
D051017:Apoptosis Regulatory Proteins; D061945:Caspase Inhibitors; D004791:Enzyme Inhibitors; D015852:Eukaryotic Initiation Factor-2; D020672:I-kappa B Proteins; D019409:Interleukin-15; D008055:Lipids; C086724:Lipofectamine; D008562:Membrane Glycoproteins; D016328:NF-kappa B; D012330:RNA, Double-Stranded; D053221:TNF-Related Apoptosis-Inducing Ligand; D014409:Tumor Necrosis Factor-alpha; D019014:fas Receptor; D000072000:NF-KappaB Inhibitor alpha; D009569:Nitric Oxide; D019892:eIF-2 Kinase; D053454:Caspase 12; D053148:Caspase 3; D053181:Caspase 8; D053453:Caspase 9; D020169:Caspases; D011070:Poly I-C