Subsets of T and natural killer (NK) lymphocytes express the CD94‐NKG2A heterodimer, a receptor for major histocompatibility complex class I molecules. We show here that engagement of the CD94‐NKG2A heterodimer inhibits both antigen‐driven tumor necrosis factor (TNF) release and cytotoxicity on melanoma‐specific human T cell clones. Similarly, CD16‐mediated NK cell cytotoxicity is extinguished by cross‐linking of the CD94‐NKG2A heterodimer. Combining in vivo and in vitro analysis, we report that both I/VxYxxL immunoreceptor tyrosine‐based inhibition motifs (ITIM) present in the NKG2A intracytoplasmic domain associate upon tyrosine phosphorylation with the protein tyrosine phosphatases SHP‐1 and SHP‐2, but not with the polyinositol phosphatase SHIP. Determination of the dissociation constant, using surface plasmon resonance analysis, indicates that NKG2A phospho‐ITIM interact directly with the SH2 domains of SHP‐1 and SHP‐2 with a high affinity. Engagement of the CD94‐NKG2A heterodimer therefore appears as a protein‐tyrosine phosphatase‐based strategy that negatively regulates both antigen‐induced T cell response and antibody‐induced NK cell cytotoxicity. Our results suggest that this inhibitory pathway sets the threshold of T and NK cell activation.