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. 2011 Sep;31(18):3802–3819. doi: 10.1128/MCB.01368-10

Fig. 7.

Fig. 7.

Association of DDX60 with RLRs. (A to C) Vectors expressing HA-tagged DDX60 were transfected into HEK293FT cells with FLAG-tagged LGP2, RIG-I, MDA5, IPS-1, IKK-ε, and/or Ubc13, and cell lysates were prepared. The lysates were treated with RNase III (B) or RNase A (C). Immunoprecipitation was carried out with anti-FLAG antibody, and the precipitates (IP) and 10% of whole-cell extract (WCE) were analyzed using SDS-PAGE. Proteins were stained by Western blotting using anti-HA or anti-FLAG antibody. (D and E) HEK293FT cells were transfected with empty or HA-tagged DDX60-expressing vectors, and cells were stimulated with dsRNA or infected with VSV. Cell lysates were prepared at the indicated times, and immunoprecipitation was performed with anti-HA antibody. The precipitates were analyzed using SDS-PAGE, and Western blotting was carried out using anti-HA and anti-HMGB1 (D) or anti-RIG-I (E) antibodies. (F and G) Vectors expressing HA-tagged DDX60 and FLAG-tagged RIG-I (F) or MDA5 (G) were transfected into HeLa cells. After 24 h, cells were fixed and stained with anti-HA or anti-FLAG antibody and then observed using confocal microscopy. (H) The upper panel shows a schematic diagram of RIG-I partial fragments. The lower panel shows results of an immunoprecipitation assay performed as described for panel A. DDX60 was found to bind to the RIG-IC region.