3.

Determination of free Mg²⁺ concentrations [Mg²⁺]_m in isolated mitochondria dependent on [Mg²⁺]_e. Mitochondria isolated from clones MPL-O (with empty vector; red curve) and MPL-E1 (expressing shRNA-E; blue curve) after 10 days of doxycy-cline treatment were loaded with the Mg²⁺-sensitive dye mag-fura 2. Changes in free Mg²⁺ concentrations [Mg²⁺]_m over time were determined upon raising extramitochondrial Mg²⁺ concentrations [Mg²⁺]_e from nominally 0 mM to 1 mM. (For details of methods cf. legend to Supplementary Fig. 3 and [5]).