The laminin-binding integrin α3β1 is highly expressed in epidermal keratinocytes, where it regulates both cell-autonomous and paracrine functions that promote wound healing and skin tumorigenesis. However, the roles for α3β1 in regulating gene expression programs that control the behaviors of immortalized or transformed keratinocytes remain underexplored. In the current study, we used a microarray approach to identify genes that are regulated by α3β1 in immortalized keratinocytes. α3β1-Responsive genes included several genes that are involved in extracellular matrix proteolysis or remodeling, including fibulin-2 and secreted protein acidic and rich in cysteine. However, α3β1-dependent induction of specific target genes was influenced by the genetic lesion that triggered immortalization, as α3β1-dependent fibulin-2 expression occurred in cells immortalized by either SV40 large T antigen or p53-null mutation, whereas α3β1-dependent expression of secreted protein acidic and rich in cysteine occurred only in the former cells. Interestingly, quantitative PCR arrays did not reveal strong patterns of α3β1-dependent gene expression in freshly isolated primary keratinocytes, suggesting that this regulation is acquired during immortalization. p53-null keratinocytes transformed with oncogenic RasV12 retained α3β1-dependent fibulin-2 expression, and RNAi-mediated knockdown of fibulin-2 in these cells reduced invasion, although not their tumorigenic potential. These findings demonstrate a prominent role for α3β1 in immortalized/transformed keratinocytes in regulating fibulin-2 and other genes that promote matrix remodeling and invasion.