Autocrine motility factor (AMF)/phosphoglucose isomerase (PGI) is a ubiquitous cytosolic enzyme and is produced as a leaderless secretory protein, released from cells via a nonclassical pathway. AMF/PGI acts extracellularly as a potent mitogen/cytokine (CXXC, chemokine). Increased expression of AMF/PGI and its receptor/CXXC-R has been found in a wide spectrum of malignancies, and is associated with cancer progression and metastasis. To directly elucidate the functional role of AMF/PGI on cell motility and neoplastic transformation, we stably transfected AMF/PGI cDNA into NIH-3T3 cells. Ectopic overexpression of AMF/PGI results in its secretion and activation via a constitutive autocrine activation loop that renders the cells highly motile, acquiring a transformed phenotype in vitro and tumorigenicity in vivo. The transformed phenotype of AMF/PGI-transfected cells leads in part resistance to induction of apoptosis induced by serum starvation, through the activation of phosphatidylinositol 3'-kinase/Akt signaling pathway and down-regulation of caveolin-1 expression. Overexpression of this housekeeping gene induces resistance to apoptosis and neoplastic transformation, and, thus, AMF/PGI represents a novel class of oncogenic protein.