Regulation of nucleocytoplasmic trafficking of transcription factor OREBP/TonEBP/NFAT5.

This article describes for the first time which specific regions of the tonicity enhancer-binding protein (TonEBP)/NFAT5 transcription factor are essential for its transport from the cytoplasm to the nucleus under hypertonic conditions and nuclear export under hypotonic conditions. The article is significant since it maps the different regions of the TonEBP/NFAT5 protein to specific trafficking mechanisms. The manuscript illustrates how molecular tools can be used to analyze complex cellular trafficking mechanisms. Understanding these mechanisms is necessary to develop better drugs for patients suffering from diabetes and immune-mediated diseases.

The transcription factor TonEBP/NFAT5 consists of an N-terminal transactivation domain, a bipartite nuclear localization signal (NLS), a Rel-homologous DNA binding domain and a glutamine-rich transactivation domain at the C-terminus. Using flag-tagged deletion constructs of the TonEBP protein transfected into HeLa cells, the authors examined which part of the TonEBP is necessary for nuclear import and export. They found that the C-terminal domain is not required for nucleocytoplasmic trafficking in response to changes in tonicity. In additional experiments using a mutant TonEBP/NFAT5 lacking activation domain 2 (AD2), they found that the first 581 amino acid residues of TonEBP/NFAT5 contain essential protein domains required for nucleocytoplasmic trafficking. Large proteins shuttle between their nucleus and the cytoplasm through nuclear pore complexes by virtue of their NLS and nuclear export signal (NES), which are recognized by specific import and export receptors. Using mutant constructs of the TonEBP/NFAT5 NLS regions, the results indicate that the NLS in TonEBP contributes to the nuclear import. To identify the functional domain responsible for hypotonicity-induced nuclear export, the authors generated deletion mutants of FLAG-TonEBP/NFAT5 and determined the subcellular localization of these mutants under different tonicities. Their data demonstrate a putative auxiliary export domain that is critical for the hypotonicity-induced nuclear export of TonEBP/NFAT5, and the data show that this export mechanism is mediated through binding to the nuclear receptor CRM1. TonEBP/NFAT5 is a ubiquitous transcription factor expressed in kidney cells, heart cells, T cells, neurons, keratinocytes, HeLa cells and many other mammalian cells. TonEBP/NFAT5 target genes are involved in osmoregulation, organ development and in the cellular immune response. Regulation of nuclear trafficking mechanisms of TonEBP/NFAT5 might play an important role in controlling transcription of its target genes.