Induction of type I interferon by RNA viruses: cellular receptors and their substrates

Baum, Alina; García-Sastre, Adolfo

The same biochemical studies, however, also highlight the fact that while ATP hydrolysis is required for RIG-I signaling it is not sufficient as a large number of RNA molecules are capable of inducing ATPase activity in vitro while failing to induce IFN production upon transfection into cells (Schlee et al. 2009; Schmidt et al. 2009; Takahasi et al. 2008) . A critical role of the RD domain in ATP hydrolysis has also been demonstrated, as the helicase domain alone possesses much lower ATPase activity in vitro in the presence of in vitro transcribed (IVT) RNA or synthetic dsRNA than RD with helicase domain . The role of the helicase/translocase function of RIG-I remains poorly understood. It is not clear whether RIG-I unwinds dsRNA duplexes in vivo or simply translocates on the RNA molecule, leaving it intact. Like all characterized helicases, RIG-I is capable of unwinding dsRNA in vitro (Takahasi et al. 2008) . However, the rate of helicase activity of RIG-I in vitro inversely correlated with the immunostimulatory potential of the RNA substrate. As RNA molecules which induced highest helicase activity possessed a 3 0 overhang or were complexed with DNA in a heteroduplex, it is difficult to ascertain the relationship of these types of molecules to the viral lifecycle. On the other hand, the lack of unwinding by IFN inducing dsRNA could indicate that RIG-I does not unwind dsRNA in vivo but simply moves along it. Translocation activity of RIG is reported in a study by Myong et al. which illustrated that RIG-I movement on dsRNA does not involve unwinding of the RNA duplex (Myong et al. 2009 ). The same study also found that the rate of translocation and ATPase activity by full length RIG-I is much higher on 5 0 ppp containing RNA than on dsRNA with a 5 0 OH group. The rate of ATP hydrolysis and translocation was similar between full length RIG-I on 5 0 ppp containing RNA with RIG-I-DCARD on synthetic dsRNA, implying that the displacement of CARD by 5 0 ppp binding of the RD allows for more rapid RIG-I movement and associated ATPase activity. Examination of whether change in ssRNA length or dsRNA length had an effect on translocation rate indicated that RIG-I translocates on the dsRNA portion of the molecule. It is important to keep in mind that under infection conditions the RNA molecules recognized by RIG-I are likely to be complexed with nucleoprotein in RNP structures. The demonstrated ability of many helicases to displace protein from RNP complexes during movement (Jankowsky and Fairman 2007) could provide an interesting mechanism for RIG-I substrate recognition where upon binding to any exposed dsRNA the helicase could proceed to move along the dsRNA and displace nucleoprotein until a 5 0 ppp group was found at which time the CARDs would be displaced and signaling could initiate.