An increased expression of the integrin α6β4 is correlated with a poor prognosis in patients with squamous cell carcinomas. However, little is known about the role of α6β4 in the early stages of tumor development. We have isolated cells from mouse skin (mouse tumor-initiating cells [mTICs]) that are deficient in both p53 and Smad4 and carry conditional alleles of the β4 gene (Itgb4). The mTICs display many features of multipotent epidermal stem cells and produce well-differentiated tumors after subcutaneous injection into nude mice. Deletion of Itgb4 led to enhanced tumor growth, indicating that α6β4 mediates a tumor-suppressive effect. Reconstitution experiments with β4-chimeras showed that this effect is not dependent on ligation of α6β4 to laminin-5, but on the recruitment by this integrin of the cytoskeletal linker protein plectin to the plasma membrane. Depletion of plectin, like that of β4, led to increased tumor growth. In contrast, when mTICs had been further transformed with oncogenic Ras, α6β4 stimulated tumor growth, as previously observed in human squamous neoplasms. Expression of different effector-loop mutants of Ras^V12 suggests that this effect depends on a strong activation of the Erk pathway. Together, these data show that depending on the mutations involved, α6β4 can either mediate an adhesion-independent tumor-suppressive effect or act as a tumor promotor.