Rac1 localizes to the mitochondrial membranes. (A) Jurkat and CEM cells overexpressing Bcl-2 (30 × 10⁶) were lysed and fractionated into purified heavy membrane (HM) and cytosolic (S) fractions. The fractions were then probed for VDAC1, Bcl-2, Rac1, and Cu/Zn SOD. (B) Rat cell lines overexpressing Bcl-2 targeted to the mitochondria (30 × 10⁶) were lysed and fractionated into purified heavy membrane (HM) and cytosolic (S) fractions. The fractions were then probed for VDAC1, Bcl-2, and Rac1. (C) Representative electron micrographs of HeLa/Neo and HeLa/Bcl-2 cells. The cells were stained with anti-Rac1 and anti–Bcl-2 antibodies and prepared for electron microscopy as described in “Immunochemical staining and transmission electron microscopy.” Arrows point to immunoreactive regions on the mitochondria showing the presence of Rac1 and Bcl-2 on the inner and outer mitochondrial membranes of HeLa/Neo and HeLa/Bcl-2. Bar represents 1 μm. Data presented are representative of at least 3 independent experiments.