Fig. 1.

ES supplementation rescues CcO assembly defects by restoring mitochondrial copper levels of coa6Δ cells. (A) Serially diluted WT and coa6Δ cells were seeded on the indicated plates and incubated at 30 °C and 37 °C for 2 d (YPD) or 4 d (YPGE) before imaging. (B–F) WT, coa6Δ and coa6Δ cells supplemented with 20 nM ES were cultured in YP galactose medium until early stationary growth phase followed by (B) oxygen consumption rate (OCR) measurement, (C) BN-PAGE/Western analysis of mitochondrial respiratory chain supercomplexes containing Complex IV (CIV; also called CcO), (D) quantification of supercomplexes, (E) in-gel activity staining for Complex IV, and (F) quantification of CcO activity, (G) mitochondrial copper levels, and (H) total cellular copper content. Error bars represent mean ± SD (n = 3, two-tailed unpaired Student’s t test, *P < 0.05 and **P < 0.005). Data shown in A, C, and E are representative of at least three independent experiments.