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Figure 4 WA352 interacts with the nuclear-encoded mitochondrial protein COX11. (a) Interaction of WA352 with COX11 proteins of rice and A. thalianaby Y2H assay and mapping of WA352 regions (shaded) for the interaction. The combination of WA352 with the empty prey vector pGADT7, and that of the empty bait vector pGBKT7 with OsCOX11, served as negative controls. The yeast cells were grown on SD/-Leu/-Trp/-His/-Ade medium. TM, predicted transmembrane segment. aa, amino acids. (b) Y2H assay mapping of the OsCOX11 region (shaded) interacting with WA352. (c) The WA352-interacting domain in OsCOX11 and the conserved sequence in A. thalianaCOX11 (AtCOX11). (d) Rice protoplasts were co-transformed with four constructs that expressed, respectively, MTS-mOrange (a red fluorescent protein variant) for marking mitochondria, OsCOX11-CFP (cyan fluorescent protein) for mitochondrial localization of OsCOX11, and MTS-SPYNE-WA352218–352and MTSSPYCE-OsCOX11 for a BiFC assay of the in vivoWA352-OsCOX11 interaction. Scale bars, 10 µm.