Although generally acknowledged as a plasma membrane protein, the epidermal growth factor (EGF) receptor has been found in the nucleus and subcellular organelles. Recently, the mitochondrial localization of the EGF receptor (EGFR) was reported; nevertheless, the molecular mechanism underlying EGFR localization in mitochondria is largely unknown. Using immunofluorescence and immunoelectron microscopy, we observed that EGFR did localize within mitochondria. Moreover, EGFR mitochondrial translocation can be increased by rapamycin treatment in A431 cells and greatly reduced by the presence of 3-methyladenine (3-MA), an inhibitor of autophagy. The reduction of mitochondrial EGFR via autophagy inhibition is further confirmed by small interference RNA (siRNA), through which the essential protein Beclin 1 was depleted. Knocking down Beclin 1 markedly decreased the mitochondrial translocation of EGFR that was induced by rapamycin. We also noticed that the content of mitochondrial EGFR transfer is decreased when the cells are exposed to the apoptotic inducer etoposide. Additionally, either EGF treatment or EGFR knockdown by siRNA results in a greater decline of cell viability in cells possessing more mitochondrial EGFRs. Taken together, we conclude that EGFR mitochondrial localization is regulated by either autophagy or programmed cell death and is correlated with cell survival.