Gabriel CH, Gross F, Karl M, Stephanowitz H, Hennig AF, Weber M, Gryzik S, Bachmann I, Hecklau K, Wienands J, Schuchhardt J, Herzel H, Radbruch A, Krause E, Baumgrass R

Transcription factors of the nuclear factor of activated T cell (NFAT)-family are essential for antigen-specific T cell activation and differentiation. Their cooperative DNA binding with other transcription factors, such as AP1-proteins (FOS, JUN, JUNB), FOXP3, IRFs and EGR1, dictate the gene regulatory action of NFATs. To identify as yet unknown ...

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interaction partners of NFAT, we purified biotin tagged NFATc1/αA, NFATc1/βC and NFATc2/C protein-complexes and analyzed their components by SILAC-based mass spectrometry. We revealed more than 170 NFAT associated proteins, half of which are involved in transcriptional regulation. Among them are known, as well as many unknown interaction partners of NFATc1 and NFATc2 in T cells, such as Raptor, CHEK1, CREB1, RUNX1, SATB1, Ikaros and Helios. The association of NFATc2 with several other transcription factors is DNA-dependent, indicating cooperative DNA binding. Moreover, our computational analysis discovered that binding motifs for RUNX and CREB1 are found preferentially in the direct vicinity of NFAT binding motifs and in a distinct orientation to them. Furthermore, we provide evidence that mTOR and CHEK1 kinase activity influence NFAT's transcriptional potency. Finally, our dataset of NFAT-associated proteins provides a good basis to further study NFAT's diverse functions and how these are modulated due to the interplay of multiple interaction partners.

Date: Sep. 16, 2016

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