There is accumulating evidence for CD5 as an important regulator of antigen receptor mediated activation in T and B cells. However, the underlying signal transduction mechanisms are still unclear. A yeast two hybrid screen using the 94 aa tail of CD5 as "bait" led to the isolation of the regulatory subunit of CK2, β. The constitutive association between CK2 and CD5 was confirmed by co-immunoprecipitation experiments in murine B lymphomas (CH12 and NYC31), human T leukemia cell line (Jurkat) and murine splenocytes. Deletion analysis of CD5 cytoplasmic tail established that all the interaction involved the CK2 sites S459 and S461, and site specific mutation of these two sites resulted in complete loss of phosphorylation by CK2. The specific association of CD5 with the regulatory subunit, β, of CK2 led us to ask if the enzyme was regulated by CD5. In B and T cell lines, cross linking of CD5 leads to activation of both CD5 associated and cytosolic CK2. In thymocytes, CD5 crosslinking resulted in activation of CD5 associated CK2, but in contrast to that observed in transformed cells, cytosolic CK2 was inhibited. In spleen cells, CD5 crosslinking did not affect CD5 associated CK2 activity but inhibited cytosolic CK2. Since CK2 is a major regulator of several signal transduction and growth associated pathways, regulation of its activity will have direct effect on multiple pathways. The association and direct regulation of CK2 with a cell surface receptor are novel observations which suggest a signal transduction pathway yet undescribed for a lymphocyte receptor.