Inhibition of T cell signaling by immunophilin-ligand complexes correlates with loss of calcineurin phosphatase activity

Abstract

Calcineurin, a Ca2+, calmodulin-dependent protein phosphatase, was recently found to bind with high affinity to two different immunosuppressant binding proteins (immunophilins) with absolute dependence on the presence of the immunosuppressants FK506 or cyclosporin A (CsA) [Liu et al. (1991) Cell 66, 807-815]. The binding affinities of the immunophilin-drug complexes toward calcineurin and the stoichiometry of the resultant multimeric complexes have now been determined, and structural elements of FK506, CsA, and calcineurin that are critical for mediating their interactions have been identified. Analogues of FK506 (FK520, FK523, 15-O-demethyl-FK520) and CsA (MeBm2t1-CsA and MeAla6-CsA) whose affinities for their cognate immunophilins do not correlate with their immunosuppressive activities have been prepared and evaluated in biochemical and cellular assays. We demonstrate a strong correlation between the ability of these analogues, when bound to their immunophilins, to inhibit the phosphatase activity of calcineurin and their ability to inhibit transcriptional activation by NF-AT, a T cell specific transcription factor that regulates IL-2 gene synthesis in human T cells. In addition, FKBP-FK506 and CyP-CsA do not inhibit members of the PP1, PP2A, and PP2C classes of serine/threonine phosphatases. These data suggest that calcineurin is the relevant cellular target of these immunosuppressive agents and is involved in Ca(2+)-dependent signal transduction pathways in, among others, T cells and mast cells.

Signaling Events Leading to Crassulacean Acid Metabolism Induction in the Common Ice Plant

“…The requirement of Ca 2ϩ for CAM signal transduction presents the possibility that Ca 2ϩ -dependent PP activities participate in signaling events leading to CAM induction. To test the possible involvement of calcineurin in signaling CAM induction, detached leaves were treated with cyclosporin A (CsA), which forms cyclophilin-CsA complexes that inhibit Ca 2ϩ /CaM-activated PP2B (Liu et al, 1992), prior to stress or ABA treatments. CsA stimulated Ppc1 transcript accumulation in unstressed (control) leaves and caused a slight super-accumulation of transcripts in leaves exposed to dehydration stress (Fig.…”

Dynamic equilibrium between calcineurin and kinase activities regulates the phosphorylation state and localization of the nuclear factor of activated T-cells

“…The ability of a number of FK506 and CsA analogues to inhibit CaN phosphatase activity is correlated with their ability to inhibit T-cell activation, consistent with the hypothesis that CaN is a cellular target of the drugs. In addition, overexpression of a constitutively active form of CaN augments T-cell activation resulting from stimulation by both Ca# + and protein kinase C (PKC), suggesting that CaN has a key role in the physiological pathway of T-cell activation [11][12][13].…”

NMR identification of calcineurin B residues affected by binding of a calcineurin A peptide

“…In CHAPS-CnB, the N-terminal residues (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14), the C-terminal residues (158-169) and the linker connecting the two domains (residues 81-86) show increased mobility as judged by narrow, intense ~SN-~H cross-peaks and relatively fast amide proton exchange [15,27]. In CnB-P2465, only the N-terminus and the last three C-terminal residues retain this increased mobility.…”