Transcription factors belonging to the nuclear factor of activated T cells 
(NFAT) family regulate the expression of cytokine genes and other inducible 
genes during the immune response. The functions of NFAT proteins are directly 
controlled by the calcium- and calmodulin-dependent phosphatase calcineurin. 
Here we show that the binding of calcineurin to NFAT is substantially increased 
when calcineurin is activated with calmodulin and calcium. FK506.FKBP12 
drug-immunophilin complexes inhibited the interaction of NFAT with activated 
calcineurin much more effectively than they inhibited the interaction with 
inactive calcineurin, suggesting that part of the interaction with activated 
calcineurin involved the enzyme active site. We have previously shown that NFAT 
is targeted to inactive calcineurin at a region distinct from the calcineurin 
active site (Aramburu, J., Garcia-Cozar, F. J., Raghavan, A., Okamura, H., Rao, 
A., and Hogan, P. G. (1998) Mol. Cell 1, 627-637); this region is also involved 
in NFAT binding to activated calcineurin, since binding is inhibited by an NFAT 
peptide spanning the calcineurin docking site on NFAT. The interacting surfaces 
are located on the catalytic domain of the calcineurin A chain and on an 
86-amino acid fragment of the NFAT regulatory domain. NFAT binding to the 
calcineurin catalytic domain was inhibited by the calcineurin autoinhibitory 
domain and the RII substrate peptide, which bind in the calcineurin active site, 
as well as by the NFAT docking site peptide, which binds to a region of 
calcineurin distinct from the active site. We propose that, in resting cells, 
NFAT is targeted to a region of the calcineurin catalytic domain that does not 
overlap the calcineurin active site. Upon cell activation, displacement of the 
autoinhibitory domain by calmodulin binding allows NFAT to bind additionally to 
the calcineurin active site, thus positioning NFAT for immediate 
dephosphorylation at functional phosphoserine residues.