NF-ATp, a T lymphocyte DNA-binding protein that is a target for calcineurin and immunosuppressive drugs.
NF-ATp, a T lymphocyte DNA-binding protein that is a target for calcineurin and immunosuppressive drugs.
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DOI：
10.1016/s0021-9258(18)53757-1
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发表时间：
1993-02
期刊：
影响因子：
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通讯作者：
P. Mccaffrey;B. Perrino;T. Soderling;A. Rao
中科院分区：
文献类型：
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作者：
P. Mccaffrey;B. Perrino;T. Soderling;A. Rao
活化T细胞核因子（NF-AT）是T细胞活化后白细胞介素-2基因转录所必需的。在这里，我们使用一种技术，涉及从SDS-丙烯酰胺凝胶洗脱和复性的蛋白质，以确定一个DNA结合的组成部分NF-AT（NF-ATP），是目前在低渗提取物的T细胞活化前，并出现在核提取物时，T细胞被激活。NF-ATp主要以表观分子量为110，000 - 140，000的迁移形式存在于静息T细胞中，而来自活化T细胞的核提取物的NF-ATp以较低的表观分子量（90，000 - 125，000）迁移。这种差异可能反映了NF-ATp的去磷酸化，因为在体外用小牛肠磷酸酶或钙和钙调蛋白依赖性磷酸酶钙调神经磷酸酶处理NF-ATp会导致其表观分子量的类似降低。我们表明，NF-ATP是去磷酸化的细胞裂解液中的钙依赖性的过程中，包括EGTA或钙调磷酸酶的特异性肽抑制剂在细胞裂解缓冲液中被阻断。此外，细胞提取物中NF-AT β的去磷酸化通过用免疫抑制药物环孢菌素A或FK 506预先处理T细胞而被抑制，所述免疫抑制药物环孢菌素A或FK 506在与它们的特异性结合蛋白亲环素和FK 506结合蛋白复合时抑制钙调磷酸酶的磷酸酶活性。这项工作确定NF-ATp作为DNA结合磷蛋白和药物/免疫亲和素/钙调神经磷酸酶复合物的目标，认为介导抑制白细胞介素-2基因诱导环孢菌素A和FK 506。
The nuclear factor of activated T cells (NF-AT) is essential for transcription of the interleukin-2 gene upon T cell activation. Here we use a technique involving elution and renaturation of proteins from SDS-acrylamide gels to identify a DNA-binding component of NF-AT (NF-ATp) that is present in hypotonic extracts of T cells prior to activation and appears in nuclear extracts when T cells are activated. NF-ATp is present in resting T cells predominantly in a form migrating with an apparent molecular weight of 110,000-140,000, while NF-ATp from nuclear extracts of activated T cells migrates with a lower apparent molecular weight (90,000-125,000). This difference is likely to reflect dephosphorylation of NF-ATp, since treatment of NF-ATp with calf intestinal phosphatase or the calcium- and calmodulin-dependent phosphatase calcineurin in vitro results in a similar decrease in its apparent molecular weight. We show that NF-ATp is dephosphorylated in cell lysates by a calcium-dependent process that is blocked by inclusion of EGTA or a specific peptide inhibitor of calcineurin in the cell lysis buffer. Moreover, dephosphorylation of NF-ATp in cell extracts is inhibited by prior treatment of T cells with the immunosuppressive drugs cyclosporin A or FK506, which inhibit the phosphatase activity of calcineurin when complexed with their specific binding proteins, cyclophilin and FK506-binding protein. This work identifies NF-ATp as a DNA-binding phosphoprotein and a target for the drug/immunophilin/calcineurin complexes thought to mediate the inhibition of interleukin-2 gene induction by cyclosporin A and FK506.