Figure 4.

ots1 ots2 Mutants Are Defective in deSUMOlyation of SUMO1/2 Conjugates.

(A) Immunoblot (IB) analysis of total protein (10 μg loaded for each lane) derived from wild-type Col-0 Arabidopsis seedlings grown for 12 d under long-day conditions in the presence of the indicated NaCl concentrations. Filters were probed with anti-SUMO1/2 (anti At S1/2) antibodies. The vertical bar shows the laddering effect produced by increased accumulation of SUMO1/2 conjugates. The arrowhead indicates free (nonconjugated) SUMO1/2. The bottom panels show immunostaining of UGPase and Ponceau staining of ribulose-1,5-bis-phosphate carboxylase/oxygenase (Rubisco) small subunit (RbscS), which served as loading controls. The experiment was repeated three times with similar results.

(B) Immunoblots of total proteins (10 μg loaded for each lane) derived from wild-type Col-0 Arabidopsis seedlings and the indicated genotypes grown in the absence (−) or presence (+) of supplemented 100 mM NaCl for 12 d under long-day conditions. Top, the filter was probed with anti-SUMO1/2 (anti At S1/2) antibodies. The arrowhead and vertical bar indicate free SUMO1/2 and conjugated SUMO1/2, respectively. The experiment was repeated three times with similar results. Bottom, the filter was probed with anti-SUMO3 (anti At S3) antibodies. UGPase immunostaining and Ponceau staining of Rubisco small subunit (RbscS) were used as loading controls.