Phosphorylation within the MafA N Terminus Regulates C-terminal Dimerization and DNA Binding
Phosphorylation within the MafA N Terminus Regulates C-terminal Dimerization and DNA Binding
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DOI：
10.1074/jbc.m110.105759
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发表时间：
2010-04-23
影响因子：
4.8
通讯作者：
Stein, Roland
中科院分区：
文献类型：
--
作者：
Guo, Shuangli;Vanderford, Nathan L.;Stein, Roland
磷酸化通过影响二聚化、细胞定位、激活电位和/或DNA结合来调节转录因子活性。然而，人们对这种翻译后修饰是如何介导这些过程的还知之甚少。在这里，我们研究了磷酸化对MafA和MafB的dna结合特性的作用，MafA和MafB是与细胞分化和肿瘤发生相关的碱性亮氨酸拉链（b-Zip）家族密切相关的转录激活因子。质谱法鉴定了许多常见的磷酸化位点。然而，去磷酸化仅阻止了对MafA二聚体的检测，因此显著降低了dna结合能力。对MafA/B嵌合体的分析表明，对MafA磷酸化状态的敏感性是由跨越c端二聚化区域（氨基酸（aa） 279-359）的序列赋予的，而同源的MafB区域（aa 257-323）传递磷酸化不依赖的DNA结合。突变分析表明，能够结合DNA的MafA二聚体的形成需要在不同的n端反激活域（aa 1-72）磷酸化，而不是在c端b-Zip区域磷酸化。这些结果表明富磷氨基酸的转激活与b-Zip结构域在控制MafA dna结合活性方面存在新的关系。
Phosphorylation regulates transcription factor activity by influencing dimerization, cellular localization, activation potential, and/or DNA binding. Nevertheless, precisely how this post-translation modification mediates these processes is poorly understood. Here, we examined the role of phosphorylation on the DNA-binding properties of MafA and MafB, closely related transcriptional activators of the basic-leucine zipper (b-Zip) family associated with cell differentiation and oncogenesis. Many common phosphorylation sites were identified by mass spectrometry. However, dephosphorylation only precluded the detection of MafA dimers and consequently dramatically reduced DNA-binding ability. Analysis of MafA/B chimeras revealed that sensitivity to the phosphorylation status of MafA was imparted by sequences spanning the C-terminal dimerization region (amino acids (aa) 279-359), whereas the homologous MafB region (aa 257-323) conveyed phosphorylation-independent DNA binding. Mutational analysis showed that formation of MafA dimers capable of DNA binding required phosphorylation within the distinct N-terminal transactivation domain (aa 1-72) and not the C-terminal b-Zip region. These results demonstrate a novel relationship between the phosphoamino acid-rich transactivation and b-Zip domains in controlling MafA DNA-binding activity.