Staphylococcus aureus is an important human and animal pathogen. During infection, this bacterium is able to attach to and enter host cells by using its cell surface-associated factors to bind to the host's extracellular matrix (ECM) proteins. In this study, we determined that a protein exported by S.aureus, α-toxin, can interfere with the integrin-mediated adhesion and internalization of S.aureus by human lung epithelial cells (A549). The downregulation of α-toxin production significantly increased bacterial adhesion and invasion into the epithelial cells. In contrast, bacterial adhesion and invasion was inhibited by both overproduction of α-toxin and the addition of α-toxin to the culture medium. Moreover, our results showed that the quantitative effects on invasion closely parallel those of adherence. This suggests that the effect on invasion is probably secondary to, and a consequence of, the reduced adherence caused by α-toxin exposure. Specifically, we demonstrated that α-toxin interacts with the hosts' ECM protein's receptor, β1-integrin, which indicates that β1-integrin may be a potential receptor of α-toxin on epithelial cells. Taken together, our results indicate that exported α-toxin inhibits the adhesion and internalization of S. aureus by interfering with integrin-mediated pathogen-host cell interactions.