FIG 3.

Hsp72 was upregulated and recruited to the IBs during MuV infection. (A) Summary of Hsp70 family protein candidates for binding partners of the P protein. (B) FLAG-Hsp70 proteins were coexpressed with HA-P in 293T cells, immunoprecipitated (IP) with anti-HA antibody, and immunoblotted (IB) with anti-HA and anti-FLAG antibodies. (C) Vero cells infected with MuV were immunostained at 24 h p.i. with mouse anti-Hsp72 or anti-Hsc70 MAb and rabbit anti-MuV V/P PAb or mouse anti-P MAb and rabbit anti-GRP78 PAb, followed by the appropriate Alexa-conjugated secondary antibodies. The cell nuclei were stained with DAPI (blue). (D) Vero cells were infected with MuV at a multiplicity of infection (MOI) of 1.0. The cell lysates were collected at the indicated times and subjected to immunoblotting with the appropriate antibodies. (E) 293T cells were transfected with pCAGGS-P (P) or empty vector (EV). At 24 h posttransfection, the cell lysates were collected and subjected to immunoblotting with the appropriate antibodies. (F and G) Vero cells transfected with pCAGGS-P alone (F) or pCAGGS-N and -P (G) were immunostained at 24 h posttransfection with mouse anti-Hsp72 MAb and rabbit anti-V/P PAb, followed by AF488-conjugated anti-mouse IgG and AF594-conjugated anti-rabbit IgG. The cell nuclei were stained with DAPI (blue).