Targeted inhibition of Zika virus infection in human cells by CRISPR-Cas13b.

Zika virus (ZIKV) outbreaks occurred in recent years on an unprecedented scale, which caused fever and severe complications like Guillain-Barré syndrome in adults and fetal abnormalities. No vaccines or other effective treatments against ZIKV are available to date. The CRISPR-Cas13 family has the unique ability to target single-strand RNA molecules and mediate RNA cleavage. In the present study, we sought to exploit CRISPR-Cas13b for developing an anti-ZIKV system in mammalian cells. We first generated a ZIKV infection and reporting system by: (1) fusing mCherry to the ZIKV capsid protein for reporting infection by fluorescence; and (2) deriving a 293T cell line (293T-DC-SIGN) stably expressing DC-SIGN (Dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin) that became highly susceptible to ZIKV infection. The CRISPR Cas13b expression was reported to be in the cytoplasm of 293T-DC-SIGN cells using a Cas13b-GFP fusion expression vector. Fourteen CRISPR RNAs (crRNAs) were designed to target the most conserved regions of the ZIKV genome through bioinformatics analysis of 1138 ZIKV genome sequences. Five crRNAs were found to have significant effects (p < 0.001; two-sided t test) for Cas13b-targeted inhibition on ZIKV infection in 293T-DC-SIGN cells. Our study demonstrated an exciting example of using the CRISPR-Cas13b system for the treatment and prevention of ZIKV infection, highlighting CRISPR-Cas13 as a promising therapeutic anti-RNA virus strategy.