Crk-associated substrate lymphocyte type (Cas-L) is a docking protein that is heavily tyrosine phosphorylated by the engagement of beta1 integrins in T cells. In the present study, we attempted to evaluate the role of Cas-L in the pathophysiology of adult T-cell leukemia (ATL). Examination of peripheral blood mononuclear cells from ATL patients as well as ATL-derived T cell lines showed an elevation of Cas-L in these cells. We showed that tyrosine phosphorylation as well as expression of Cas-L was markedly elevated through the induction of human T-lymphotropic virus type I (HTLV-I) Tax in JPX-9 cells, with these cells showing marked motile behavior on the ligands for integrins. We next performed yeast two-hybrid screening of cDNA library from an HTLV-I-transformed T cell line, which resulted in the identification of Tax as a putative binding partner for Cas-L. Co-precipitation experiments revealed that the serine-rich region of Cas-L might serve as the binding site with the highest affinity for Tax. Co-localization study showed that Tax and Cas-L partly merged in the cytoplasm. Finally, we showed that exogenous Cas-L inhibited Tax-mediated transactivation of nuclear factor kappaB (NF-kappaB), while Tax-independent activation of NF-kappaB remained intact, hence indicating that Cas-L might specifically regulate Tax-NF-kappaB pathway.