Contribution of IPS-1 to polyI:C-induced cytokine production in conjunctival epithelial cells.
We previously demonstrated that ocular surface epithelium expressed TLR3 and that its ligand, polyI:C, stimulation induced the secretion of inflammatory cytokines and type I IFN. It was recently reported that RIG-I and MDA5 also recognize viral dsRNA mimicking polyI:C. In this study, we investigated whether RIG-I and/or MDA5 contribute to polyI:C-inducible responses in conjunctival epithelium. The expression of RIG-I, MDA5, and TLR3 in human conjunctival epithelium was examined by RT-PCR and their up-regulation after polyI:C stimulation by quantitative RT-PCR and immunoblot analysis. Human conjunctival epithelial cells also expressed RIG-I, MDA5 and TLR3 mRNA and protein. The expression of RIG-I and MDA5, but not of TLR3, was markedly up-regulated upon polyI:C stimulation. We also examined the function of IPS-1 (an adaptor molecule common to RIG-I and/or MDA5) and TLR3 in conjunctival epithelium using IPS-1 KO and TLR3 KO mice. To analyze in vivo murine conjunctival epithelial cells, 10 μl of a 100 μg/ml polyI:C solution were delivered subconjunctivally and as eye drops, then conjunctival epithelial cells were subjected to gene expression analysis. We focused on 10 transcripts up-regulated in murine conjunctival epithelium upon polyI:C stimulation. Cxcl10, Mx1, Ifi44, Ifi203, Iigp2 and Rtp4 were dominantly regulated by IPS-1, Ccl5 by TLR3, and Rsad2, Mx2 and Cmpk2 were regulated by TLR3 and IPS-1. Our results showed that conjunctival epithelial cells express RIG-I and MDA5, and IPS-1, an adaptor molecule common to RIG-I and MDA5, contributes to polyI:C-inducible cytokine production in conjunctival epithelial cells.
Department of Ophthalmology, Kyoto Prefectural University of Medicine, Kyoto, Japan. mueta@koto.kpu-m.ac.jp
Mayumi
M
Ueta
Taro
T
Kawai
Norihiko
N
Yokoi
Shizuo
S
Akira
Shigeru
S
Kinoshita
D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D003228:Conjunctiva / Q000276:immunology*; D016207:Cytokines / Q000096:biosynthesis*; D000071457:DEAD Box Protein 58; D053487:DEAD-box RNA Helicases / Q000235:genetics / Q000378:metabolism; D004847:Epithelial Cells / Q000187:drug effects / Q000276:immunology*; D004848:Epithelium / Q000187:drug effects / Q000276:immunology; D015870:Gene Expression; D006801:Humans; D000072640:Interferon-Induced Helicase, IFIH1; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism; D011070:Poly I-C / Q000276:immunology* / Q000494:pharmacology; D011956:Receptors, Cell Surface; D011971:Receptors, Immunologic; D051196:Toll-Like Receptor 3 / Q000235:genetics / Q000378:metabolism; D015854:Up-Regulation; D000097905:Roundabout Proteins
D048868:Adaptor Proteins, Signal Transducing; D016207:Cytokines; D000071457:DEAD Box Protein 58; D053487:DEAD-box RNA Helicases; D000072640:Interferon-Induced Helicase, IFIH1; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D011070:Poly I-C; D011956:Receptors, Cell Surface; D011971:Receptors, Immunologic; D051196:Toll-Like Receptor 3; C505371:MAVS protein, human; C118348:Robo3 protein, mouse; D000097905:Roundabout Proteins; C452820:RIGI protein, human; C452815:IFIH1 protein, human; C510592:Ifih1 protein, mouse; C512612:MAVS protein, mouse