The ubiquitously expressed c-Abl tyrosine kinase localizes to the nucleus and cytoplasm. Using confocal microscopy, we demonstrated that c-Abl colocalizes with the endoplasmic reticulum (ER)-associated protein grp78. Expression of c-Abl in the ER was confirmed by immunoelectron microscopy. Subcellular fractionation studies further indicate that over 20% of cellular c-Abl is detectable in the ER. The results also demonstrate that induction of ER stress with calcium ionophore A23187, brefeldin A, or tunicamycin is associated with translocation of ER-associated c-Abl to mitochondria. In concert with targeting of c-Abl to mitochondria, cytochrome c is released in the response to ER stress by a c-Abl-dependent mechanism, and ER stress-induced apoptosis is attenuated in c-Abl-deficient cells. These findings indicate that c-Abl is involved in signaling from the ER to mitochondria and thereby the apoptotic response to ER stress.