Inhibition of calcineurin phosphatase activity by a calcineurin B homologous protein

Abstract

Calcineurin, a Ca2+/calmodulin-stimulated protein phosphatase, plays a key role in T-cell activation by regulating the activity of NFAT (nuclear factor of activated T cells), a family of transcription factors required for the synthesis of several cytokine genes. Calcineurin is the target of the immunosuppressive drugs cyclosporin A and FK506 complexed with their cytoplasmic receptors cyclophilin and FKBP12, respectively. In this study we report that calcineurin is also the target of a recently identified Ca2+- binding protein, CHP (for calcineurin homologous protein), which shares a high degree of homology with the regulatory B subunit of calcineurin and with calmodulin. In Jurkat and HeLa cells, overexpression of CHP specifically impaired the nuclear translocation and transcriptional activity of NFAT but had no effect on AP-1 transcriptional activity and only a small (<25%) inhibitory effect on the transcriptional activity of NFκB. Further study indicated that CHP inhibits calcineurin activity. In cells overexpressing CHP, the phosphatase activity of immunoprecipitated calcineurin was inhibited by ~50%; and in a reconstituted assay, the activity of purified calcineurin was inhibited up to 97% by the addition of purified recombinant CHP in a dose-dependent manner. Moreover, prolonged activation of Jurkat cells was associated with a decreased abundance of CHP, suggesting a possible regulatory mechanism allowing activation of calcineurin. CHP, therefore, is a previously unrecognized endogenous inhibitor of calcineurin activity.