Program of Cellular Biology, Brazilian National Cancer Institute (INCA), Rio de Janeiro, Brazil.
Cell Cycle. 2012 Mar 1;11(5):1014-28. doi: 10.4161/cc.11.5.19518.
A number of physiological processes in both normal and cancer cells are regulated by the proto-oncogene c-Myc. Among them, processes such as cell cycle regulation, apoptosis, angiogenesis and metastasis are also controlled by the nuclear factor of activated T cells (NFAT) family of transcription factors. It is already known that NFAT upregulates c-Myc expression by binding to an element located in the minimal c-Myc promoter. However, the importance of other NFAT sites in the context of the full promoter has not been evaluated. In this work, we demonstrate that the regulation of c-Myc by NFAT1 is more complex than previously conceived. In addition to the proximal site, NFAT1 directly binds to distal sites in the c-Myc promoter with different affinities. Promoter deletions and site-directed mutagenesis of NFAT binding sites in HEK293T cells suggest that in NFAT1-mediated transactivation, some NFAT elements are negative and dominant and others are positive and recessive. Furthermore, we demonstrate that cooperation with partner proteins, such as p300, enhances NFAT1-mediated transactivation of the c-Myc promoter. At last, the newly identified sites are also responsive to NFAT2 in HEK293T cells. However, in NIH3T3 cells, the regulation mediated by NFAT proteins is not dependent on the known NFAT sites, including the site previously described. Thus, our data suggest that the contribution of NFAT to the regulation of c-Myc expression may depend on a balance between the binding to positive and negative NFAT-responsive elements and cooperation with transcriptional cofactors, which may differ according to the context and/or cell type.
许多正常细胞和癌细胞中的生理过程受原癌基因 c-Myc 调控。其中，细胞周期调控、细胞凋亡、血管生成和转移等过程也受激活 T 细胞核因子（NFAT）家族转录因子的调控。已知 NFAT 通过结合位于最小 c-Myc 启动子中的元件来上调 c-Myc 表达。然而，在完整启动子背景下，其他 NFAT 位点的重要性尚未得到评估。在这项工作中，我们证明 NFAT1 对 c-Myc 的调控比之前想象的更为复杂。除了近端位点外，NFAT1 还以不同亲和力直接结合 c-Myc 启动子中的远端位点。在 HEK293T 细胞中进行的启动子缺失和 NFAT 结合位点的定点突变实验表明，在 NFAT1 介导的反式激活中，一些 NFAT 元件是阴性和显性的，而另一些是阳性和隐性的。此外，我们证明与伙伴蛋白（如 p300）的合作增强了 NFAT1 介导的 c-Myc 启动子的反式激活。最后，新鉴定的位点在 HEK293T 细胞中也对 NFAT2 有反应。然而，在 NIH3T3 细胞中，NFAT 蛋白介导的调控不依赖于已知的 NFAT 位点，包括之前描述的位点。因此，我们的数据表明，NFAT 对 c-Myc 表达的调控作用可能取决于与阳性和阴性 NFAT 反应元件的结合以及与转录共因子的合作之间的平衡，这可能因上下文和/或细胞类型而异。
