Fasciculation and elongation protein zeta-1 (FEZ1) promotes efficiently the neurite elongation of rat phaeochromocytoma PC12 cells. We here characterized FEZ1 in PC12 cells. Nerve growth factor (NGF) stimulation induces significant expression of endogenous FEZ1 in PC12 cells. Upon NGF stimulation FEZ1 localizes in both cytoplasm and neuritis, co-localizing with mitochondria. Silencing of FEZ1 by RNA interference efficiently reduces NGF-induced neurite elongation and the anterograde motility of mitochondria in PC12 cells. Immunoprecipitation and pulldown assay shows that FEZ1 interacts with kinesin superfamily protein 5 (KIF5) and tubulin. Thus, our results suggest that the FEZ1/kinesin complex functions for the transport of mitochondria along microtubules toward the extending neurites in differentiating PC12 cells.