One signaling pathway known to act downstream of VEGF is regulated by the nuclear factor of activated T-cell (NFAT) family of transcription factors.
10–12 The NFAT family is a set of five proteins grouped for their similarity to the Rel Homology Domain (RHD)/nuclear factor kappa B (NFκB) family of transcription factors.
13 Within this family, NFATc refers to the four isoforms (NFATc1, NFATc2, NFATc3, and NFATc4) regulated by intracellular Ca
2+ through the serine phosphatase calcineurin (CN).
14 Calcineurin regulation of NFATc is controlled through its binding to a conserved Ca
2+/CN-dependent translocation (CAT) regulatory domain, consisting of a 300 amino acid region located at the N-terminus of the DNA binding domain, and encoded by a single exon in all four proteins. When static, the NFAT regulatory domain is heavily phosphorylated, but when CN is activated it dephosphorylates the domain revealing a nuclear localization sequence, and NFAT translocates to the nucleus where it begins to accumulate, demonstrating an increased affinity for its target DNA sites.
15,16 Efficient dephosphorylation of NFAT has been shown to require a direct docking interaction with CN, and the protein is quickly rephosphorylated by NFAT kinases in the nucleus when CN is blocked, emphasizing its dependence and sensitivity to CN activity and Ca
2+ dynamics.
17,18 Due to the tight association between CN and NFAT activity, the compound inhibitor of NFAT-calcineurin association-6 (INCA-6) is commonly used in experimental conditions to inhibit NFAT signaling.
19 -21 Inhibitor of NFAT-calcineurin association-6 is a small organic molecule (9,10-dihydro-9,10[1′,2′]-benzenoanthracene-1,4-dione) that competitively binds to the discrete NFAT binding site of CN, blocking NFAT dephosphorylation without altering other CN phosphatase activity.
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