Figure 5.

Aberrant interaction between PHB2 and COX in SphK2-null mitochondria. A–C) Analysis of mitochondrial electron transport chain complexes by BN-PAGE. A, B) Equal amounts of heart mitochondrial proteins from wild-type and sphk2^−/− mice were separated by BN-PAGE (A) or transferred to nitrocellulose and immunoblotted with the indicated specific complex antibodies (B). C) In-gel catalytic activity assay for COX using the same gel as in A. C I–V, mitochondrial complexes I–V, respectively. Arrows indicate the additional band present only in sphk2^−/− mitochondria. D, E) Interaction between PHB2 and COX. Mitochondria extracts from wild-type and sphk2^−/− hearts were immunoprecipitated with anti-PHB2 antibody (D) or with an antibody against the holocytochrome-c oxidase complex (E) and then analyzed by Western blotting with the indicated antibodies. Inputs are shown at left. Results are representative of 3 experiments.