Fig. 1.

SUMOylation of Kaiso protein under normal conditions. a Cells Caki-1, HEK293, COS-7 and two Kaiso knockout clones were lysed in loading buffer and analyzed by SDS-PAGE and western blotting, and probed with anti-Kaiso antibody to detect Kaiso and its post-translationally modified forms. Kaiso (>100 kDa) is post-translationally modified under normal conditions. b Nuclear extracts were prepared with or without SUMO isopeptidase inhibitor NEM from HEK293 (left panel) and from HEK293 transfected with Kaiso-GFP (right panel) and analyzed by SDS-PAGE, western blotting, and stained with anti-Kaiso antibody. In the presence of NEM, an additional band was detected. c The whole-cell lysates were immunoprecipitated with anti-Kaiso, anti-SUMO1, anti-SUMO2/3 with or without NEM. Western blot analysis revealed that Kaiso and SUMO1 co-immunoprecipitated. Kaiso is SUMOyaled by SUMO1. d, e Kaiso-GFP was transfected into HEK293 cells alone or with SUMO1, 2, 3,-GFP plasmids. Cells were lysed in loading buffer or immunoprecipitated with Kaiso antibodies with or without NEM and analyzed by western blot. Kaiso may be monoSUMOylated by all forms of SUMO