The E3 ligase TRIM56 is a host restriction factor of Zika virus and depends on its RNA-binding activity but not miRNA regulation, for antiviral function

Yang, Darong; Li, Nan L.; Wei, Dahai; Liu, Baoming; Guo, Fang; Elbahesh, Husni; Zhang, Yunzhi; Zhou, Zhi; Chen, Guo-Yun; Li, Kui

A subset of TRIM proteins has been shown to modulate innate immune signaling during viral infections [51] . Flavivirus replication generates dsRNAs, a major pathogen-associated molecular pattern that are sensed by two classes of pattern recognition receptors (PRRs), i.e., the RIG-I-like receptors (RLRs, RIG-I and MDA5) and TLR3 [52] . While TRIM56 is dispensable for RLR signaling and its overexpression per se does not trigger IFN production, it promotes TLR3-dependent innate immune response via interacting with the adaptor protein TRIF, independent of its E3 ligase activity [34] . It should be noted, however, that several lines of evidence suggest that the antiviral effect of TRIM56 against ZIKV revealed in this study is direct and not secondary to a regulation of IFN antiviral responses. First, ZIKV, like other flaviviruses, is not a strong inducer of IFN responses. Flaviviruses replicate their RNAs within re- TRIM56 is an RNA-binding protein that restricts Zika virus arranged membrane structures that shield and delay viral dsRNAs from innate immune recognition [53] . ZIKV encodes multiple IFN antagonists including NS1, NS4A, NS5, among others, that inhibit IFN production and/or signaling [54] [55] [56] . Consistent with this, we found almost no induction of IFN-β, IFN-λ1 and ISGs by ZIKV in cell culture within 48 h.p.i (S7 Fig). Second, TRIM56 did not augment, but rather decreased, the minimal induction of IFNs and ISG56 late post ZIKV infection (S7 Fig) , which may be explained by TRIM56-mediated reduction in ZIKV RNA (i.e., the immune stimulus) levels. Third, although ZIKV may elicit TLR3 signaling, as suggested by a recent study showing that ZIKV could disrupt neurogenesis through TLR3 activation [57] , and TRIM56 promotes TLR3 signaling [34] , the E3 ligase-deficient TRIM56 mutant (AA) with intact function in augmenting TLR3/TRIF signaling [34] was incapable of suppressing ZIKV propagation. Furthermore, we demonstrated that efficient knockdown of TLR3 by siRNA ( S8A Fig) had no impact on the anti-ZIKV activity of TRIM56 in HEK293 cells (S8B Fig). Altogether, the anti-ZIKV action of TRIM56 is not attributed to its effect on TLR3 signaling. At present, we cannot fully exclude the possibility that in certain cell types such as immune cell subsets, TRIM56 may promote innate immune activation and finetune host responses, thereby adding to its direct antiviral effect. Whether this mechanism operates will require future studies.