UC Riverside

The Flavivirus genus consists of over 70 vector-borne, single-stranded RNAcontaining

viruses, including dengue virus (DENV) and Zika virus (ZIKV), which cause

major epidemics among humans and pose a serious threat to global public health. No

vaccines or antivirals exist to prevent or treat infections caused by DENV and ZIKV. To

establish infection, it is vital for flaviviruses to overcome the antiviral state induced by

type 1 interferon. ZIKV and DENV NS5 proteins can bind to human signal transducer

and activator of transcription 2 (hSTAT2) protein, a key signaling intermediary for

interferon (IFN) responses, which triggers the subsequent proteasome-dependent

degradation of hSTAT2. NS5 does not have a host counterpart; thus, NS5 or the NS5-

hSTAT2 interaction are logical antiviral targets. However, the underlying mechanism of

the Flavivirus NS5-hSTAT2 interaction remains elusive. Here, we have elucidated the

structure of the ZIKV NS5 protein and re-purposed a small molecule inhibitor against

DENV for the inhibition of ZIKV RdRp activity. We have also delineated the biochemical and structural basis of the DENV and ZIKV NS5-hSTAT2 interactions and used the knowledge gained from those studies to generate hSTAT2 and NS5 mutants

deficient in interaction. We discovered that the mechanism of hSTAT2 interaction is

conserved among the DENV2 and ZIKV NS5 proteins. In addition to hSTAT2

degradation, we found that ZIKV NS5 also competes with IRF9 for hSTAT2 binding,

resulting in a two-pronged ISGF3 suppression mechanism. Finally, the hSTAT2-

interaction deficient NS5 mutants were used as tools to interrogate the functional

consequences of the hSTAT2-NS5 interaction during ZIKV infection. Disruption of the

ZIKV NS5-hSTAT2 interaction resulted in loss of NS5-mediated hSTAT2 degradation

and IFN suppression. Importantly, during infection, this interaction is critical for efficient

ZIKV propagation. The work described here provides critical insights into DENV2 and

ZIKV NS5-mediated hSTAT2 degradation and suppression of the IFN response and will

facilitate the development of novel vaccines and inhibitors of Flavivirus infections.