Regulation and function of the cytosolic viral RNA sensor RIG-I in pancreatic beta cells.
Enteroviral infections are associated with type I diabetes. The mechanisms by which viruses or viral products such as double-stranded RNA (dsRNA) affect pancreatic beta cell function and survival remain unclear. We have shown that extracellular dsRNA induces beta cell death via Toll-like receptor-3 (TLR3) signaling whereas cytosolic dsRNA triggers the production of type I interferons and apoptosis via a TLR3-independent process. We presently examined expression of the intracellular viral RNA sensors, the RNA helicases RIG-I and MDA5, and documented the functionality of RIG-I in pancreatic beta cells. FACS-purified rat beta cells and islet cells from wild-type or TLR3(-/-) mice were cultured with or without the RIG-I-specific ligand 5'-triphosphate single-stranded RNA (5'triP-ssRNA), the synthetic dsRNA polyI:C (PIC) or 5'OH-ssRNA (negative control); the RNA compounds were added in the medium or transfected in the cells using lipofectamine. RIG-I and MDA5 expression were determined by real-time RT-PCR. NF-kappaB and IFN-beta promoter activation were studied in the presence or absence of a dominant-negative form of RIG-I (DN-RIG-I). Both extracellular (PICex) and intracellular (PICin) PIC increased expression of RIG-I and MDA5 in pancreatic beta cells. TLR3 deletion abolished PICex-induced up-regulation of the helicases in beta cells but not in dendritic cells. PICin-induced NF-kappaB and IFN-beta promoter activation were prevented by the DN-RIG-I. The RIG-I-specific ligand 5'triP-ssRNA induced IFN-beta promoter activation and beta cell apoptosis. Our results suggest that the RIG-I pathway is present and active in beta cells and could contribute to the induction of insulitis by viral RNA intermediates.
García
Mónica
M
Laboratory of Experimental Medicine, Université Libre de Bruxelles, Brussels, Belgium.
Dogusan
Zeynep
Z
Moore
Fabrice
F
Sato
Shintaro
S
Hartmann
Gunther
G
Eizirik
Decio L
DL
Rasschaert
Joanne
J
D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000235:genetics; D003600:Cytosol / Q000201:enzymology* / Q000821:virology; D053487:DEAD-box RNA Helicases / Q000096:biosynthesis; D004770:Enterovirus / Q000378:metabolism*; D004769:Enterovirus Infections / Q000201:enzymology* / Q000235:genetics; D017353:Gene Deletion; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects / Q000235:genetics; D050417:Insulin-Secreting Cells / Q000201:enzymology* / Q000821:virology; D000072640:Interferon-Induced Helicase, IFIH1; D016899:Interferon-beta / Q000096:biosynthesis; D008297:Male; D008565:Membrane Proteins / Q000096:biosynthesis*; D051379:Mice; D018345:Mice, Knockout; D016328:NF-kappa B / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000096:biosynthesis*; D011070:Poly I-C / Q000494:pharmacology; D011401:Promoter Regions, Genetic / Q000235:genetics; D012330:RNA, Double-Stranded / Q000378:metabolism*; D012367:RNA, Viral / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar; D011956:Receptors, Cell Surface; D051196:Toll-Like Receptor 3 / Q000235:genetics / Q000378:metabolism; D015854:Up-Regulation / Q000187:drug effects / Q000235:genetics; D000097905:Roundabout Proteins
D053487:DEAD-box RNA Helicases; D000072640:Interferon-Induced Helicase, IFIH1; D016899:Interferon-beta; D008565:Membrane Proteins; D016328:NF-kappa B; D009419:Nerve Tissue Proteins; D011070:Poly I-C; D012330:RNA, Double-Stranded; D012367:RNA, Viral; D011956:Receptors, Cell Surface; D051196:Toll-Like Receptor 3; C118348:Robo3 protein, mouse; D000097905:Roundabout Proteins; C519703:TLR3 protein, mouse; C530472:TLR3 protein, rat; C510592:Ifih1 protein, mouse