Figure 4
From: Foot-and-mouth disease virus non-structural protein 3A inhibits the interferon-β signaling pathway
FMDV 3A and its mutants interact with RIG-I, MDA5, VISA and their mutants.
(a–c) A schematic diagram of full-length and truncated FMDV 3A, RIG-I, MDA5 and VISA and their mutants. (d–f) Amino acids 1–51 of 3A are required for its interaction with RIG-I, MDA5 and VISA. The HEK293T cells (2 × 106) were cotransfected with 4 μg of FMDV GFP-3A plasmid or its mutants and the plasmids expressing Flag-RIG-I (F-RIG-I), Flag-MDA5 (F-MDA5) or Flag-VISA (F-VISA) (5 μg each). Coimmunoprecipitations were performed with anti-Flag (F) or control IgG (lg) antibodies. Immunoblot analysis was performed with anti-green fluorescent protein (GFP) (upper panels). Expression levels of the proteins were analyzed by immunoblot analysis of the lysates with antibodies against anti-Flag and anti-GFP (lower panels). (g,h) Amino acids 360–540 of VISA and the CARD and helicase domains of RIG-I/MDA5 interacts with 3A. The HEK293T cells (2 × 106) were co-transfected with full-length or truncated HA-RIG-I (10 μg), HA-MDA5 (10 μg) or HA-VISA (4 μg) and Flag-3A (4 μg) (F-3A). Coimmunoprecipitations were performed with anti-HA (H), anti-Flag (F) or control IgG (lg) antibodies. Immunoblot analysis was performed with antibodies against anti-Flag-HRP or anti-HA. Expression levels of the proteins were analyzed by immunoblot analysis of the lysates with anti-HA and anti-Flag (lower panels). HC: heavy chain; LC: light chain.
