H omeobox transcription factors specify body plan by regulating differentiation, proliferation, and migration at a cellular level. The homeobox transcription factor Gax is expressed in quiescent vascular smooth muscle cells (VSMCs), and its expression is downregulated by vascular injury or other conditions that lead to VSMC proliferation. Previous investigations demonstrate that Gax may regulate VSMC proliferation by upregulating the cyclin-dependent kinase (cdk) inhibitor p21. Here we examined whether Gax influences VSMC migration, a key feature in the development of stenotic lesions after balloon injury. Transduction of a Gax cDNA inhibited the migratory response of VSMCs toward PDGF-BB, basic fibroblast growth factor, or hepatocyte growth factor/scatter factor. Gax expression also inhibited migration of NIH·3T3 fibroblasts and embryonic fibroblasts lacking p53. Gax was unable to inhibit the migration of fibroblasts lacking p21, but this effect could be restored in these cells by providing exogenous p21 or by overexpressing another cdk inhibitor, p16. Flow cytometric analysis implicated a Gax-mediated downregulation of α_vβ₃ and α_vβ₅ integrin expression in VSMCs as a potential cause for reduced cell motility. Gax specifically downregulated β₃ and β₅ in VSMCs in culture and after acute vascular injury in vivo. Repression of integrin expression was also found in NIH 3T3 cells and p53 knockout fibroblasts, but not in p21-knockout fibroblasts, unless these cells express exogenous p21 or p16. These data suggest that cycle progression, integrin expression, and cell migration can be regulated in VSMCs by the homeobox gene product Gax.J. Clin. Invest. 104:1469–1480 (1999).