Cell surface proteins of the transmembrane 4 superfamily (TM4SF) are a newly characterized family of proteins which are presumed to span the plasma membrane four times. The function of this family of molecules is poorly understood, but based on monoclonal antibody studies there is some evidence that they may be involved in transmembrane signal transduction and regulation of cell proliferation, differentiation, or both, in a number of different cell types. CD53 is a member of this family that is expressed on leukocytes, and transduces activation signals through unknown mechanisms that may involve phosphorylation events. However, CD53 has never been shown to associate directly with kinases. Here, we show by immunoprecipitation from cell lysates of lymph nodes and a thymoma cell line, that immune complexes of rat CD53 contain tyrosine phosphatase activity. The CD53-associated phosphatase was able to dephosphorylate in vitro the phosphorylated tyrosine kinase Lck, as well as a synthetic substrate, and its activity was abrogated by a tyrosine phosphatase inhibitor. Although its identity has not been established, it is clear from depletion experiments that it is not CD45. CD63, a second member of the TM4SF, also co-precipitates a phosphatase activity from rat basophilic leukemia cells. These results demonstrate that the TM4SF members associate with tyrosine phosphatases. It seems possible that such associated phosphatases may contribute to the signal transduction capacity of TM4SF molecules.