SIM plays a role in the recruitment of Sp1 into the PML NBs.

<p>A. Wild-type PML, PML3 m, PMLas, and PML3mas expression plasmids were transfected into PML^−/− MEFs separately. Total proteins were isolated from different plasmid transfected cells and co-immunoprecipitations using anti-PML antibody were performed, * in the left panel represents a negative control with anti IgG, and western blotting was carried out with anti-Sp1 antibody. Western blotting on 10% of protein used for immunoprecipitation was performed for inputs detection. B. Double-color immunofluorescence staining with Sp1 polyclonal and PML monoclonal antibodies was performed in PML^−/− MEFs transfected with different plasmids described in A. DNA were counterstained with DAPI. C. The Sp1 SUMOylation site is essential for physical interaction with PML. PML^−/− MEFs were co-transfected by PMLIV with pCMV-FLAG-Sp1-HA or pCMV-FLAG-Sp1K16R-HA. Immunoprecipitation was performed with anti-PML antibody, and Western blotting was performed with anti-HA antibody. * in the right panel represents a negative control with anti IgG, Western blotting on 10% of protein used for immunoprecipitation was performed for inputs detection. D. Double-color immunofluorescence staining with HA antibody and PML monoclonal antibodies was performed in the transfected cells as described in C, followed by confocal microscopic analysis.