Subcellular localization of 6K in infected cells.

(A) Schematic of the miniSOG-6K SINV construct. The sequence encoding the fluorescent protein miniSOG was cloned into WT toto64 after Thr2 of 6K as an N-terminal fusion. (B) Western blot analysis of the cell lysates from BHK-15 cells infected with WT SINV or miniSOG-6K SINV. The blot was probed with an anti-E2 primary antibody to detect the structural polyprotein processing intermediates. (C-E) Representative images of BHK-15 cells transfected with mCherry-SEC61B-C1 (red) (C), mTagBFP2-SiT-N-15 (blue) (D), or CP-E3-mCherry-E2-6K/TF-E1 (red) (E), and infected with miniSOG-6K SINV (green). Imaging was performed at 12 hpi. Regions of interest are marked and enlarged to the right as separate channels. Images correspond to videos (S3–S5 Videos) (F-G) Representative images of BHK-15 cells electroporated with RNA corresponding to the dual-labeled mCherry-E2/miniSOG SINV (mCherry-E2 is red and miniSOG-6K is green) (F) or ΔTF mCherry-E2/miniSOG SINV (mCherry-E2 is red and miniSOG-6K is green) (G) and imaged at 12 hpi Images correspond to videos (S6 and S7 Videos).