Fig. 2. IPI7 ubiquitinates IPA1.

a In vitro ubiquitination of His-IPA1 by MBP-IPI7. MBP-IPI7(H58Y) and MBP were used as negative controls. Immunoblotting was performed with antibodies against His, MBP or Ub. The presence (+) or absence (−) of components in the reaction mixture was indicated. Similar results were obtained from three independent biological experiments. b IPI7 promotes ubiquitination of IPA1 in vivo. The assay was performed with rice variety Ri22 and Ri22-derived transgenic plants overexpressing IPI7 driven by a rice ubiquitin promoter (IPI7-OE). Immunoprecipitation was performed with an IPA1 antibody and immunoblotting was performed with an antibody against Ub or IPA1. Similar results were obtained from three independent biological experiments.