Figure 2

Subcellular localization of N protein together with all viral components. (a) Vero E6 cells were transfected with pCAGGS-SFTSV-N, pCAGGS-SFTSV-GP, pCAGGS-SFTSV-RdRp, and vRNA-Rluc and subsequently fixed at 24 hpt. The cells were stained with antibodies against N protein or GP (green) and ERGIC or Golgi organelle markers (red), and the nuclei were stained with DAPI (blue). The yellow areas in the merged images show the cellular localization of proteins with organelle markers. Magnification of the merged area is shown on the right side of the merged image. The mock-transfected cells are shown as mock images. (b) Localization of N, ERGIC, and Golgi apparatus shown in (a) was analyzed using ImageJ (corresponding to * P < 0.05, ** P < 0.01, and *** P < 0.001). Number of cells = n. (c) Detection of transcriptional activity using a minigenome assay. BHK/T7-9 cells were transfected with pCAGGS-SFTSV-N, pCAGGS-SFTSV-RdRp, and vRNA-Rluc or pCAGGS-SFTSV-N, pCAGGS-SFTSV-RdRp, vRNA-Rluc, and pCAGGS-SFTSV-GP. The luciferase activities were compared with that of N + vRNA-Rluc (without RdRp) as a negative control. The activity of Rluc was standardized by Fluc activity (transfection control). Results are shown as fold increases (RLU Rluc/Fluc) in the bar graph with standard deviations of the means. The statistical significance between RdRp and without RdRp in different combinations was assessed using the Student’s t-test (corresponding to *P < 0.05 and ** P < 0.01).