Fig. 5.

Supercomplex assembly during ERS. Abundance of ETC supercomplexes is increased by ERS. Mitochondria were prepared from wild-type (WT), pex34Δ and cta1Δ cells treated with or without tunicamycin (0.5 µg/ml; tun) for 5 h, and from cells with or without constitutive expression of CPY*, as described in the Materials and Methods, and then frozen. Cells constitutively expressing CPY* were not treated with tunicamycin. For analysis on blue native PAGE gels, mitochondria were thawed, solubilized with digitonin and run on a 3–12% gradient gel, then transferred to PVDF membrane, as described in the Materials and Methods. Supercomplexes were analyzed by western blotting with anti-Cox2 and anti-Qcr7 antibodies. Asterisks indicate a III₂+IV₂ tetramer; circles indicate a III₂IV₁ trimer. Positions of molecular mass markers (kDa) are shown. Images are representative of three experiments.