Figure 2.
NSP12 is recruited to the NSP3/4-induced DMVs through NSP3–NSP12 interaction. (A) Confocal microscopy analysis showing the colocalization of NSP12-mCherry with Flag-NSP3/NSP4-GFP–induced punctate structures in HeLa cells. Trace outline is used for line-scan analysis of the relative fluorescence intensity of NSP12-mCherry, Flag-NSP3, and NSP4-GFP. Scale bar, 10 μm. (B and C) Confocal microscopy analysis showing the colocalization of NSP12-mCherry with GFP-NSP3 (B) or NSP4-GFP (C) in HeLa cells. Trace outline is used for line-scan analysis of the relative fluorescence intensity of NSP12-mCherry with GFP-NSP3 or NSP4-GFP. Scale bar, 10 μm. (D) Co-IP analysis of the interaction of Flag-NSP3 or NSP4-Flag with NSP12-mCherry. Proteins were transiently expressed in HEK-293T cells and immunoprecipitates pulled down by Flag antibody were analyzed by IB with the indicated antibodies. Flag-vector was used as a negative control. Input represents 5% of the total cell extract used for IP. Molecular weights are in kDa. n = 3 independent experiments. (E) Co-IP analysis of the interaction of NSP12-mCherry with Flag-NSP3 or NSP4-Flag. Proteins were transiently expressed in HEK-293T cells and immunoprecipitates pulled down by mCherry antibody were analyzed by IB with the indicated antibodies. Flag-vector was used as a negative control. Input represents 5% of the total cell extract used for IP. Molecular weights are in kDa. n = 3 independent experiments. (F) Co-IP analysis of the interaction of NSP12 with NSP3 in the SARS-CoV-2–infected Vero-E6 cells. IgG was used as a negative control. Molecular weights are in kDa. n = 3 independent experiments. Source data are available for this figure: SourceData F2.

NSP12 is recruited to the NSP3/4-induced DMVs through NSP3–NSP12 interaction. (A) Confocal microscopy analysis showing the colocalization of NSP12-mCherry with Flag-NSP3/NSP4-GFP–induced punctate structures in HeLa cells. Trace outline is used for line-scan analysis of the relative fluorescence intensity of NSP12-mCherry, Flag-NSP3, and NSP4-GFP. Scale bar, 10 μm. (B and C) Confocal microscopy analysis showing the colocalization of NSP12-mCherry with GFP-NSP3 (B) or NSP4-GFP (C) in HeLa cells. Trace outline is used for line-scan analysis of the relative fluorescence intensity of NSP12-mCherry with GFP-NSP3 or NSP4-GFP. Scale bar, 10 μm. (D) Co-IP analysis of the interaction of Flag-NSP3 or NSP4-Flag with NSP12-mCherry. Proteins were transiently expressed in HEK-293T cells and immunoprecipitates pulled down by Flag antibody were analyzed by IB with the indicated antibodies. Flag-vector was used as a negative control. Input represents 5% of the total cell extract used for IP. Molecular weights are in kDa. n = 3 independent experiments. (E) Co-IP analysis of the interaction of NSP12-mCherry with Flag-NSP3 or NSP4-Flag. Proteins were transiently expressed in HEK-293T cells and immunoprecipitates pulled down by mCherry antibody were analyzed by IB with the indicated antibodies. Flag-vector was used as a negative control. Input represents 5% of the total cell extract used for IP. Molecular weights are in kDa. n = 3 independent experiments. (F) Co-IP analysis of the interaction of NSP12 with NSP3 in the SARS-CoV-2–infected Vero-E6 cells. IgG was used as a negative control. Molecular weights are in kDa. n = 3 independent experiments. Source data are available for this figure: SourceData F2.

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