Japan. 8 These authors contributed equally to this work. Correspondence should be addressed to A.T. (takaoka@igm.hokudai.ac.jp). At least three DExD/H-box RNA helicases 4,5 , RIG-I, MDA5 and LGP2 are included in the RLR family. RIG-I is a key PRR for the detection of positive-and negative-stranded RNA viruses in the cytoplasm of cells 6,7 , and plays an important role in triggering responses against many viruses, such as orthomyxovirus (influenza A virus) and paramyxovirus (measles, mumps, and Sendai virus (SeV)) families, hepatitis C virus (HCV), and Japanese encephalitis virus (JEV) 8 . 5'-triphosphate modifications of RNA (3pRNA) are essential for RIG-I recognition and activation 9,10 . Ligand-binding activates the ATPase activity of RIG-I to change its structural conformation 6 , which in turn enables RIG-I to interact through its N-terminal tandem caspase recruitment domain (CARD) with the adaptor protein MAVS (for mitochondrial antiviral signaling protein, also known as IPS-1, VISA, or Cardif) [11][12][13][14] . MAVS then initiates the activation of interferon (IFN)-regulatory factor (IRF) 3, IRF7 and NF-κB transcriptional pathways for the subsequent production of type I IFNs and inflammatory cytokines, which are crucial for activating innate immune responses to viral infection 6,15 . Given the important role of the RIG-I pathway in the antiviral innate response, the mechanisms regulating RIG-I activation represent a topic of intense research [16][17][18] .Poly(ADP-ribose) polymerases (PARPs), a superfamily with least 17 members, are known to regulate not only cell survival and cell death programs triggered by DNA 3 damage, but also other biological functions as well as pathological processes, such as inflammatory and degenerative diseases, in a manner dependent or independent of their PARP activity [19][20][21][22] . Several PARP-superfamily members have a direct regulatory effect on replication of certain viruses 19,20,22,23
RESULTS
4PARPs contribute to the IFN response To investigate the role of the PARP-superfamily members in nucleic acid induced innate immune responses, we selected some of the PARP-superfamily members known to be involved in microbial infection, inflammation and immunity: PARP-1, PARP-2, PARP-7, PARP-9, 23,[31][32][33][34]. We then examined whether they have the ability to enhance the induction of IFNB mRNA in HEK293T cells in response to stimulation with three different types of nucleic acids-3pRNA, poly(rI:rC) and poly(dA-dT)•poly(dA-dT) (named poly(dA:dT) hereafter).Among the protein tested, PARP-13 uniquely showed a marked enhancing effect on IFNB mRNA expression induced by stimulation with 3pRNA, poly(rI:rC) and poly(dA:dT) ( Fig. 1a), all of which are known to activate the RIG-I-mediated pathway in HEK293T cells 9,10,[35][36][37] . A weak increase in IFNB mRNA expression was also detected in cells expressing PARP-1, PARP-2 and PARP-9. PARP-13 exists in at least two isoforms 22,26,38 . The amino-terminal 254-amino acid fragment of the rat homologue, which corresponds to the N...
