Figure 5.
SARS-CoV-2 N protein inhibits the binding between TAK1 and TAB3, unlike SARS-CoV N protein. (A) HEK293T cells were transfected with Flag-TAK1, Flag-SARS-CoV-2 N, Flag-empty vector, HA-TAB3, and HA-empty vector. Lysates were collected for immunoprecipitation (IP) (anti-HA) and immunoblot (IB) analysis. The ratio of the gray value between IP TAK1-Flag and IP TAB3-HA was determined. (B) HEK293T cells were transfected with Myc-empty vector or Myc-SARS-CoV-2 N for 48 h, followed by TNF-α treatment for 4 h. Lysates were subjected to immunoprecipitation (anti-TAB3) and immunoblot analysis. The ratio of the gray value between IP TAK1 and IP TAB3 was determined. (C) HEK293T cells were transfected with Flag-TAK1, Flag-empty vector, Myc-SARS-CoV N, Myc-empty vector, HA-TAB3, and HA-empty vector. Lysates were collected for immunoprecipitation (anti-HA) and immunoblot analysis. The ratio of the gray value between IP TAK1-Flag and IP TAB3-HA was determined. (D) HEK293T cells were transfected with Myc-empty vector or Myc-SARS-CoV N for 48 h, followed by TNF-α treatment for 4 h. Lysates were subjected to immunoprecipitation (anti-TAB3) and immunoblot analysis. The ratio of the gray value between IP TAK1 and IP TAB3 was determined. (E) Domain organization of TAB3 and its truncated mutants. CUE, Ubiquitin binding domain; TBD, TAK1 binding domain; NZF, NLP4-like zinc finger domain. (F–H) HEK293T cells were transfected Flag-empty vector, Flag-TAK1 (F), Flag-SARS-CoV-2 N (G), or Flag-SARS-CoV N (H) along with HA-TAB3 full-length (FL), HA-TAB2 (1–478aa), and HA-TAB2 (1–661aa). Lysates were subjected to immunoprecipitation (anti-Flag) and immunoblot analysis. Data are done at least twice, and one representative is shown. Source data are available for this figure: SourceData F5.

SARS-CoV-2 N protein inhibits the binding between TAK1 and TAB3, unlike SARS-CoV N protein. (A) HEK293T cells were transfected with Flag-TAK1, Flag-SARS-CoV-2 N, Flag-empty vector, HA-TAB3, and HA-empty vector. Lysates were collected for immunoprecipitation (IP) (anti-HA) and immunoblot (IB) analysis. The ratio of the gray value between IP TAK1-Flag and IP TAB3-HA was determined. (B) HEK293T cells were transfected with Myc-empty vector or Myc-SARS-CoV-2 N for 48 h, followed by TNF-α treatment for 4 h. Lysates were subjected to immunoprecipitation (anti-TAB3) and immunoblot analysis. The ratio of the gray value between IP TAK1 and IP TAB3 was determined. (C) HEK293T cells were transfected with Flag-TAK1, Flag-empty vector, Myc-SARS-CoV N, Myc-empty vector, HA-TAB3, and HA-empty vector. Lysates were collected for immunoprecipitation (anti-HA) and immunoblot analysis. The ratio of the gray value between IP TAK1-Flag and IP TAB3-HA was determined. (D) HEK293T cells were transfected with Myc-empty vector or Myc-SARS-CoV N for 48 h, followed by TNF-α treatment for 4 h. Lysates were subjected to immunoprecipitation (anti-TAB3) and immunoblot analysis. The ratio of the gray value between IP TAK1 and IP TAB3 was determined. (E) Domain organization of TAB3 and its truncated mutants. CUE, Ubiquitin binding domain; TBD, TAK1 binding domain; NZF, NLP4-like zinc finger domain. (F–H) HEK293T cells were transfected Flag-empty vector, Flag-TAK1 (F), Flag-SARS-CoV-2 N (G), or Flag-SARS-CoV N (H) along with HA-TAB3 full-length (FL), HA-TAB2 (1–478aa), and HA-TAB2 (1–661aa). Lysates were subjected to immunoprecipitation (anti-Flag) and immunoblot analysis. Data are done at least twice, and one representative is shown. Source data are available for this figure: SourceData F5.

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