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. Author manuscript; available in PMC: 2015 Jun 19.
Published in final edited form as: Immunity. 2014 Jun 12;40(6):936–948. doi: 10.1016/j.immuni.2014.05.007

Fig. 7. OASL specifically enhances RIG-I-signaling by mimicking pUb to activate RIG-I.

Fig. 7

(A) Ubiuitination or p(Ub) binding mutants of RIG-I can be activated in presence of OASL. Cells were transfected with wild type (Wt) or mutant RIG-I constructs for 24 h and analyzed by IB with indicated antibodies. (B) OASL is capable of promoting RIG-I activation in the absence of TRIM25. Cells were transfected with 50 nM TRIM25 siRNA (Zeng et al., 2010) or control siRNA for 48 h followed by SeV infection for 16 h and IB analysis. (C) OASL enhances RIG-I(N) oligomerizaiton similar to pUb. 50 or 100 ng of purified RIG-I(N) (GST tagged) were incubated either with K63-linked polyubiquitin chain (mixture of (pUb)>3, Boston Biochem) or with purified OASL for 1h at 30°C in the binding buffer (20 mM HEPES-KOH pH 7.4; 5 mM MgCl2 and 1X proteinase inhibitor cocktails). The incubated proteins were analyzed by native PAGE followed by IB with anti-GST antibody.