Analysis of the preexisting and nuclear forms of nuclear factor of activated T cells
Jain, J.; Miner, Z.; Rao, A.
Journal of Immunology 151(2): 837-848
1993
ISSN/ISBN: 0022-1767 PMID: 8335913 Document Number: 422263
The nuclear factor of activated T cells (NF-AT)-3 is an inducible DNA-binding protein that is essential for transcriptional induction of the IL-2 gene during T cell activation. NF-AT is thought to consist of two components: a ubiquitous, inducible nuclear component that we have identified as Fos and Jun proteins, and a preexisting, T cell-specific component (NF-AT-p) which is the target for the immunosuppressive agents cyclosporin A (CsA) and FK506. We have previously shown that nuclear extracts from activated T cells form two inducible NF-AT complexes with an oligonucleotide corresponding to the distal NF-AT site of the murine IL-2 promoter, although hypotonic extracts of unstimulated T cells form a single complex containing NF-AT-p. We show that the ability to detect NF-AT-p in a gel shift assay, which is essential for purification and biochemical studies of this protein, is strikingly dependent on the precise sequence of the NF-AT oligonucleotide used as the labeled probe. Moreover we present evidence that the component that forms the faster-migrating ("lower") nuclear NF-AT complex is derived by a calcium-dependent, cyclosporin-sensitive, posttranslational modification of NF-AT-p, and that Fos and Jun proteins stabilize its interaction with DNA. The results are discussed in the context of a model relating the two nuclear NF-AT complexes to NF-AT-p.