Fig. 4. Ubiquitylation of the Na⁺/K⁺-ATPase subunits. The HeLa cells in DMEM were incubated with or without the proteasome inhibitor lactacystin for 5 hours. The ubiquitylated proteins were immunoprecipitated from the cell lysate with anti-ubiquitin antibody (polyclonal antibody for 1, β1, ubiquitin and IgG immunoblots and monoclonal antibody for β3 immunoblot) in the presence of the inhibitor and then subjected to immunoblotting using the antibodies indicated. Mouse IgG was used in the immunoblotting as a negative control (IgG). The positions of the molecular size markers (in kDa) are indicated on the right.