Catalytic subunits of the phosphatase calcineurin interact with NF-κB-inducing kinase (NIK) and attenuate NIK-dependent gene expression

Nuclear factor (NF)-kappa B-inducing kinase (NIK) is a serine/ threonine kinase that activates NF-kappa B pathways, thereby regulating a wide variety of immune systems. Aberrant NIK activation causes tumor malignancy, suggesting a requirement for precise regulation of NIK activity. To explore novel interacting proteins of NIK, we performed in vitro virus screening and identified the catalytic subunit A alpha isoform of serine/ threonine phosphatase calcineurin (CnA alpha) as a novel NIK-interacting protein. The interaction of NIK with CnAa in living cells was confirmed by co-immunoprecipitation. Calcineurin catalytic subunit A beta isoform (CnA alpha) also bound to NIK. Experiments using domain deletion mutants suggested that CnAa and CnA beta interact with both the kinase domain and C-terminal region of NIK. Moreover, the phosphatase domain of CnA alpha is responsible for the interaction with NIK. Intriguingly, we found that TRAF(3), a critical regulator of NIK activity, also binds to CnA alpha and CnA beta. Depletion of CnA alpha and CnA beta significantly enhanced lymphotoxin-beta receptor (Lt beta R)-mediated expression of the NIK-dependent gene Spi-B and activation of RelA and RelB, suggesting that CnA alpha and CnA beta attenuate NF-kappa B activation mediated by Lt beta R-NIK signaling. Overall, these findings suggest a possible role of CnA alpha and CnA beta in modifying NIK functions.