Figure 8.

Validation of K63 Polyubiquitination for Nuclear Hits Identified by Sensor-Based Proteomics Using Transient Expression in Wild Tobacco.

(A), (C), and (E) Subcellular localization of SUN2-GFP (A), MEE5-GFP (C), and H2B.11-GFP (E) transiently expressed in N. benthamiana leaves. Bar = 10 µm.

(B), (D), and (F) In vivo K63 polyubiquitination of transiently expressed SUN2 (B), MEE5-GFP (D), and H2B.11-GFP (F). Immunoprecipitation was performed using anti-GFP antibodies on RIPA buffer-solubilized protein extracts. N. benthamiana plants expressing free GFP were used as a negative control (0). Eluates were subjected to immunoblotting with anti-GFP and anti-K63 polyUb antibodies (Apu3). The sizes of marker proteins in kilodaltons are shown. IB, immunoblotting, IP, immunoprecipitation.