Phosphorylation of NFATc4 by p38 Mitogen-Activated Protein Kinases

Abstract

Nuclear factor of activated T cells (NFAT) is implicated in multiple biological processes, including cytokine gene expression, cardiac hypertrophy, and adipocyte differentiation. A conserved NFAT homology domain is identified in all NFAT members. Dephosphorylation of the NFAT homology region is critical for NFAT nuclear translocation and transcriptional activation. Here we demonstrate that NFATc4 is phosphorylated by p38 mitogen-activated protein (MAP) kinase but not by JNK. The p38 MAP kinase phosphorylates multiple residues, including Ser¹⁶⁸ and Ser¹⁷⁰, in the NFAT homology domain of NFATc4. Replacement of Ser^168,170 with Ala promotes nuclear localization of NFATc4 and increases NFAT-mediated transcription activity. Stable expression of Ala^168,170 NFATc4, but not of wild-type NFATc4, in NIH 3T3 cells promotes adipocyte formation under differentiation conditions. Molecular analysis indicates that peroxisome proliferator-activated receptor γ2 (PPARγ2) is a target of NFAT. Two distinct NFAT binding elements are located in the PPARγ2 gene promoter. Stable expression of Ala^168,170 NFATc4, but not of wild-type NFATc4, increases the expression of PPARγ, which contributes in part to increased adipocyte formation. Thus, NFAT regulates PPARγ gene expression and has a direct role in adipocyte differentiation.

The first three authors contributed equally to this work.

We thank T. Hoey, B. M. Spiegelman, and T. Soderling for providing reagents; T. Barrett for technical assistance; and K. Gemme for administrative assistance.

This work was supported, in part, by grants from the National Cancer Institute (to R.J.D.) and the American Health Assistance Foundation (to C.-W.C.). R.J.D. is an investigator of the Howard Hughes Medical Institute.