Inhibition of cds-mediated t cell activation by fsba: evidence for an intracellular site of action

Engagement of the TCR-CD3 receptor complex by ligands activates intracellular protein tyrosine kinases (PTK) which are critical for T cell effector function Recent evidence suggests that cytolytic T lymphocyte (CTL) activity is dependent on ectoATPase activity The inhibition of ectoATPase activity and resultant loss of cytolytic activity was accomplished using 5'-p-(fluorosulfonyl)benzoyl adenosine (FSBA) an adenine nucleotide analogue. Alternatively, the loss of CTL activity may result from the ability of FSBA to inhibit intracellular PTKs required for cytolytic activity. We demonstrate that in the ectoATPase-deficient Jurkat T cell line, FSBA is capable of inhibiting intracellular PTKs. FSBA at 1 mM completely blocked Ick activity in an in vitro kinase reaction. Incubation of intact Jurkat cells with 1 mM FSBA led to the inhibition of PTK activity associated with the TCR-CD3 receptor complex following CD3 crosslinking. Consistent with these results we observed a loss of tyrosine phosphorylated substrates in CD3 crosslinked T cells treated with FSBA. Furthermore FSBA at ImM, blocked the CDS-mediated mobilization of intracellular calcium in intact Jurkat cells. These results suggest that FSBA is capable of inhibiting T cell activation via the inhibition of intracellular PTKs.