Supplementary Methods, Table 1, Figures 1-7 from Manganese Superoxide Dismutase Regulates a Metabolic Switch during the Mammalian Cell Cycle

PDF file - 964K, Results presented in this section show: (1) A minimal difference in cell cycle phase distributions in MnSOD (+/+), and (-/-) MEFs following re-entry into the proliferative cycle (Table I) (2) An increase in MnSOD protein levels and activity in MnSOD overexpressing MB231 human mammary epithelial cancer cells (Figure 1) (3) An inhibition of MnSOD activity in quiescent normal human fibroblasts was associated with an increase in glucose consumption and percent S-phase (Figure 2) (4) Cell cycle phase-associated increase in cellular ROS levels was absent in MnSOD (-/-) MEFs (Figure 3) (5) Cell cycle phase specific increase in DHE-fluorescence is primarily due to an increase in cellular steady-state levels of superoxide (Figure 4) (6) Mass spectrometry results identifying lysine and arginine methylation pattern of MnSOD during quiescence and proliferation; species conservation of lysine and arginine methylation sites in MnSOD (Figure 5) (7) Site directed mutagenesis approach to mutate lysine 89 and 202 of MnSOD, and engineer expression vectors of wild-type and K-to-A mutant carrying human MnSOD cDNAs (Figure 6). (8) Computer modeling of MnSOD-methylation pattern (Figure 7)