The integrin CD103 and the chemokine receptor CCR9 are co-expressed on small intestinal CD8(+) intraepithelial lymphocytes (IEL), naïve murine CD8(+) T cells and by a small population of effector/memory CD8(+) T cells, indicating a potential role for CCR9 in regulating CD103 expression and function. Here, we demonstrate that CD103, in contrast to CCR9, is down-regulated on CD8(+) T cells following their activation in mesenteric lymph nodes and that effector CD8(+) T cells upon initial entry into the small intestinal epithelium are CCR9(+)CD103(-). CD103 was rapidly induced on wild-type CD8(+) T cells subsequent to their entry into the small intestinal epithelium, however, CCR9(-/-) CD8(+) T cells exhibited a significant delay in CD103 induction at this site. In addition, the CCR9 ligand, CCL25, that is constitutively expressed in the small intestinal epithelium, induced transient, dose-dependent and pertussis toxin-sensitive CD103-mediated adhesion of CD8(+) small intestinal IEL to a murine E-cadherin human Fc (mEFc) fusion protein. Together, these results demonstrate a role for CCR9/CCL25 in promoting the induction and function of CD103 on CD8(+) IEL and suggest that this chemokine receptor/chemokine pair may function to regulate lymphocyte-epithelial interactions in the small intestinal mucosa.