PCV2 Cap interacts with pDNAJB6 and co-localizes in both cytoplasm and nucleus. A, B PCV2 Cap interacts with pDNAJB6 in transfected cells. HEK293T cells were co-transfected with GFP-Cap and Flag-pDNAJB6 expression plasmids or co-transfected with the pEGFP-N1 control vector and the Flag-pDNAJB6 expression plasmid. The cells transfected with pEGFP-N1 control vector, GFP-Cap expression plasmid or Flag-pDNAJB6 expression plasmid alone served as controls. The interaction of GFP-Cap with Flag-pDNAJB6 was identified through immunoprecipitation using anti-Flag antibodies or anti-GFP antibodies. C pDNAJB6 interacts with PCV2 Cap protein. PK-15 cells were infected with PCV2 (MOI = 1) for 36 h, and cell lysates were immunoprecipitated using Cap antibodies followed by immunoblots using pDNAJB6 antibodies. D Endogenous pDNAJB6 interacts with GFP-Cap. PK-15 cells were transfected with GFP-Cap, and the cells were lysed for immunoprecipitation with GFP antibodies; pDNAJB6 antibodies were used for immunoblotting. In these co-IP assay, the corresponding whole cell lysate (Lysate) were detected the targeted protein before immunoprecipitation. E Direct interaction of pDNAJB6 with PCV2 Cap. Bacterially purified GST-pDNAJB6 or GST alone was incubated with purified His-Cap, and these input proteins (Input) and the proteins pulled-down by glutathione Sepharose beads (Output) were analyzed by Western blot with the indicated antibodies. F pDNAJB6 protein co-localizes with Cap. PK-15 cells were co-transfected with GFP-Cap and Flag-pDNAJB6 expression plasmid, and transfected with pEGFP-N1 and pCI-neo expression plasmid alone served as control. Cells were fixed and then stained with DAPI (blue) at 48 h post transfection. Scale bar = 10 μm.