Mct 1

Reduced, solvent-exposed thiols on the cell surface were labeled with biotin-MTSEA (10 mM) for 15 min protected from light, resolved using non-reducing SDS-PAGE, and visualized with Streptavidin conjugate to horseradish peroxidase (HRP) [27 (link)]. Pretreatment with DTT (1 mM, 10 min) prior to labeling was used as a control for total cell-surface thiols. Biotinylated proteins were immunopreciptated with streptavidin beads and probed using antibodies specific for MCT1 and GAPDH (Cell Signaling #2118).

Cell culture reagents were purchased from ScienCell (Carlsbad, CA, USA). Antibodies against GSS, SOD, CAT, AMPK, PGC-1α, and TFAM and the goat anti-rabbit IgG and goat anti-mouse IgG antibodies were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Antibodies against ACC, HK-I, HK-II, PFK, LDHA, MCT-1 and MCT-4 were from Cell Signaling Technology (San Jose, CA, USA). Nrf-1 and Nrf-2 antibodies were purchased from Proteintech (Rosemont, IL, USA). Propidium iodide was acquired from BD bioscience (San Jose, CA, USA), and the electrophoresis reagents were from Bio-Rad (Richmond, CA, USA). The nitrocellulose membranes were purchased from Amersham Scientific (Piscataway, NJ, USA). All other reagents were purchased from Sigma–Aldrich (St. Louis, MO, USA).