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. Author manuscript; available in PMC: 2021 Jan 8.
Published in final edited form as: Nature. 2020 Jul 8;585(7825):414–419. doi: 10.1038/s41586-020-2457-8

Fig. 2. TRIM7 ubiquitinates ZIKV-E and promotes virus replication.

Fig. 2.

a, Virus titers from TRIM7 JEG-3 Knockout (TRIM7 KO) infected with ZIKVs (MOI 0.5). (n=3, technical replicates, mean +/−SE, multiple t-test, Holm-Sidak correction, ****p<0.0001). b, HEK293Ts transfected with TRIM7 or a short isoform (TRIM7 ΔRB), WT and mutant E, followed by IP. Representative of 2 independent experiments. c, TRIM7 ubiquitinates recombinant ZIKV-E on both K38 and K281 in an in vitro ubiquitination assay. Representative of four independent experiments. Densitometry shown in d (N=3, mean +/− SE, One-way Anova, Tukey’s multiple comparison, **p<0.01, ****p<0.0001, NSp>0.5).). e, K63-linked polyubiquitinated E was detected on ZIKV particles concentrated from supernatants from Vero after IP with anti-K63-linked or anti-E 4G2 antibodies. After immunoblot (IB) with anti-E, the blots were re-probed with anti-K63-Ub. f, IP with anti-E (4G2) of virus stocks from WT or TRIM7 KO JEG-3 cells. Representative from 2 independent experiments. g-i, Trim7−/− and Trim7+/+ littermates from 3 separate CRISPR KO lines (4–5 weeks old) were treated i.p. with anti-IFNAR1 (MAR1–5A3). Next day infections performed with mouse-adapted ZIKV Dakar (footpad, 105 PFU, n=7 Trim7+/+, 8 Trim7−/− mice). Serum (h) and tissue titers (i). Unpaired, t-test, two-sided *p<0.05, **p<0.01.