FIGURE 3.

In situ PLA was performed on HEK293 cells using anti-ANT2 and anti-UCP2 antibodies. Yellow dots represent a within approximately 40 nm co-localization of UCP2 and ANT2. (A) Unstimulated cells represent cells at basal condition, whereas FCCP stimulated cells represent artificially induced mitochondrial respiration and lowering of membrane potential via mitochondrial uncoupling. Oligomycin stimulated cells represent the lowest possible respiration together with the highest possible membrane potential via ATP-synthase inhibition with oligomycin. (B), Graph displaying the extent of ANT2-UCP2 interaction dependent on cellular respiration explained in (A). (C) Native PAGE was performed to confirm binding of the anti-UCP2 and anti-ANT2 to the native proteins. (D) Negative controls using only secondary antibodies or secondary antibodies in presence of either primary anti-ANT2 or anti-UCP2 antibodies. (E) Graph representing negative controls displayed as the average number of PLA signals per cell.