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Fig. 6 Mitochondrial FOXM1 decreases ETC complex protein levels and activity. (A) Western blot analysis of mitochondrial proteins. Total cellular proteins from FOXM1 stable cell lines and SH FOXM1 cells expressing MLS mutant constructs were probed for proteins from each ETC complex and β-ACTIN as a loading control. MTATP6 indicates protein subunit 6 from ETC complex V; MTCO1 indicates protein subunit 1 from ETC complex IV; UQCRC2 indicates protein subunit 2 from ETC complex III; SDHB indicates protein subunit from ETC complex II; MTND1 indicates protein subunit 1 from ETC complex I. (B) Densitometric quantification of mitochondrial proteins in stable control and OE FOXM1 and SH FOXM1 cell lines. Average MTND1, SDHB, MTCO1, UQCRC2, or MTATP6 levels were normalized to ACTIN. Values are expressed relative to Flag-FOXM1 control. Data represent mean ± SEM from three determinations. Statistical significance is indicated with an asterisk; *p < 0.05. (C) Enzymatic activities of respiratory chain complexes in stable control and OE FOXM1 and SH FOXM1 cell lines. The activities of respiratory complexes were investigated by enzymatic assays on complexes I, II, III, IV, and V in mitochondrial or total cell lysates. Activities are expressed as OD absorbance/min/mg total protein or mitochondrial protein. Data represent mean ± SEM from experiments in triplicate. Statistical significance is indicated with an asterisk; *p < 0.05. (D) Densitometric quantification of mitochondrial proteins in SH FOXM1 cells expressing MLS mutant constructs. Average MTND1, SDHB, MTCO1, UQCRC2, or MTATP6 levels were normalized to ACTIN. Values are expressed as relative to Flag-FOXM1 control. Data represent mean ± SEM from three determinations. Statistical significance is indicated with an asterisk; *p < 0.05, **p < 0.01. (E) Enzymatic activities of respiratory chain complexes in SH FOXM1 cells expressing MLS mutant constructs. The activities of respiratory complexes were investigated by enzymatic assays on complexes I, II, III, IV, and V in mitochondrial or total cell lysates. Activities are expressed as OD absorbance/min/mg total protein or mitochondrial protein. Data represent mean ± SEM from experiments in triplicate. Statistical significance is indicated with an asterisk; *p < 0.05, **p < 0.01. (F) Schematic diagram. FOXM1 translocates into the mitochondria, interacts with PTCD, and inhibits mitochondrial respiration.