Visualization and biochemical analyses of the emerging mammalian 14-3-3-phosphoproteome.
Visualization and biochemical analyses of the emerging mammalian 14-3-3-phosphoproteome.
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DOI：
10.1074/mcp.m110.005751
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发表时间：
2011-10
期刊：
影响因子：
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通讯作者：
MacKintosh C
中科院分区：
文献类型：
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作者：
Johnson C;Tinti M;Wood NT;Campbell DG;Toth R;Dubois F;Geraghty KM;Wong BH;Brown LJ;Tyler J;Gernez A;Chen S;Synowsky S;MacKintosh C
在一次描述一种相互作用的出版物以及基于高通量14-3 -3-亲和力和质谱的研究中，已经报道了数百种候选14 - 3 - 3-结合（磷酸化）蛋白。在这里，我们将这些数据转录成一种通用格式，将低通量研究的整理数据保存在MINT（http：//mint.bio.uniroma2.it/mint）中，并在易于分析和扩展的VisANT图中比较低通量和高通量数据。探索这些图提示了关于技术和生物特异性的问题，这些问题通过实验得到解决，导致鉴定了铁硫簇组装酶（ISCU）线粒体输入序列中的磷酸化14-3-3-结合位点，线粒体分裂因子（MFF）的胞质结构域，内质网束缚受体表达增强蛋白4（REEP 4），RNA调节因子SMAUG 2，和细胞骨架调节蛋白，即脱蛋白样蛋白（DBNL）和驱动蛋白轻链（KLC）同种型。因此，14-3-3s与蛋白质发生生理相互作用，这些蛋白质注定要到达不同的亚细胞位置。绘制和验证相互作用是使用14-3-3-磷酸蛋白质组学来确定健康和疾病信号通路的机制和生物标志物的基础。
Hundreds of candidate 14-3-3-binding (phospho)proteins have been reported in publications that describe one interaction at a time, as well as high-throughput 14-3-3-affinity and mass spectrometry-based studies. Here, we transcribed these data into a common format, deposited the collated data from low-throughput studies in MINT (http://mint.bio.uniroma2.it/mint), and compared the low- and high-throughput data in VisANT graphs that are easy to analyze and extend. Exploring the graphs prompted questions about technical and biological specificity, which were addressed experimentally, resulting in identification of phosphorylated 14-3-3-binding sites in the mitochondrial import sequence of the iron-sulfur cluster assembly enzyme (ISCU), cytoplasmic domains of the mitochondrial fission factor (MFF), and endoplasmic reticulum-tethered receptor expression-enhancing protein 4 (REEP4), RNA regulator SMAUG2, and cytoskeletal regulatory proteins, namely debrin-like protein (DBNL) and kinesin light chain (KLC) isoforms. Therefore, 14-3-3s undergo physiological interactions with proteins that are destined for diverse subcellular locations. Graphing and validating interactions underpins efforts to use 14-3-3-phosphoproteomics to identify mechanisms and biomarkers for signaling pathways in health and disease.