Proteomic Analysis of Apoptosis Related Proteins Regulated by Proto-Oncogene Protein DEK
Proteomic Analysis of Apoptosis Related Proteins Regulated by Proto-Oncogene Protein DEK
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DOI：
10.1002/jcb.22083
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发表时间：
2009-04-15
影响因子：
4
通讯作者：
Seo, Sang-Beom
中科院分区：
文献类型：
--
作者：
Kim, Dong-Wook;Chae, Jung-Il;Seo, Sang-Beom
核磷蛋白 DEK 与某些人类疾病有关，例如白血病和抗免疫性疾病，并且是后生动物染色质的主要成分。基本上作为染色质结构的调节剂，它可以参与各种 DNA 和 RNA 依赖性过程，并充当激活剂或阻抑剂。尽管人们做出了大量努力来表明 DEK 的生物学作用，但不同生理状态下 DEK 的直接靶蛋白仍然难以捉摸。为了研究 DEK 蛋白是否触发某些蛋白质网络的变化，使用 siRNA 表达在两种类型的细胞克隆中敲低 DEK。在这里，我们提供了来自正常 HeLa 和 DEK 敲低 HeLa 细胞的总细胞裂解物中的蛋白质组谱目录，以及用于剖析这种原癌 DEK 蛋白引起的蛋白质网络的良好体外模型系统。在此生物学背景下，我们通过二维凝胶电泳、定量图像分析和 MALDI-TOF MS 分析的组合方法比较了总蛋白质组变化。 DEK存在大量的靶标，这些靶标在DEK敲低细胞中差异表达，由58个蛋白（41个上调和17个下调）组成，通过使用特异性抗体的Western印迹分析进一步证实了一些候选亚群的差异调节表达。在已识别的 58 个点中，已知 16% 的蛋白质与细胞凋亡相关。其中，我们鉴定了细胞凋亡相关蛋白，如膜联蛋白、烯醇化酶 1、核纤层蛋白 A 和谷胱甘肽-S-转移酶 omega 1。这些结果与最近的研究一致，表明 DEK 在细胞凋亡途径中发挥着至关重要的作用。我们通过 ChIP 分析进一步证明，DEK 的敲低会导致 Prx VI 启动子周围历史的过度乙酰化，而在我们的分析中，Prx VI 启动子的表达上调。通过免疫印迹分析，我们证明了 DEK 敲低对其他 caspase 依赖性凋亡相关蛋白的调节，并进一步揭示了其在凋亡途径中的作用。 J.细胞。生物化学。 106: 1048-1059, 2009。(c) 2009 Wiley-Liss, Inc.
A nuclear phosphoprotein, DEK, is implicated in certain human diseases, such as leukemia and antoimmune disorders, and a major component of metazoan chromatin. Basically as a modulator of chromatin structure, it can involve in various DNA and RNA-dependent processes and function as either an activator or repressor. Despite of numerous efforts to suggest the biological role of DEK, direct target proteins of DEK in different physiological status remains elusive. To investigate if DEK protein triggers the changes in certain protein networks, DEK was knocked down at both types of cell clones using siRNA expression. Here we provide a catalogue of proteome profiles in total cell lysates derived from normal HeLa and DEK knock-down HeLa cells and a good in vitro model system for dissecting the protein networks due to this protooncogenic DEK protein. In this biological context, we compared total proteome changes by the combined methods of two-dimensional gel electrophoresis, quantitative image analysis and MALDI-TOF MS analysis. There were a large number of targets for DEK, which were differentially expressed in DEK knock-down cells and consisted of 58 proteins (41 up-regulated and 17 down-regulated) differentially regulated expression was further confirmed for some subsets of candidates by Western blot analysis using specific antibodies. In the identified 58 spots, 16% of proteins are known to be associated with apoptosis. Among others, we identified apoptosis related proteins such as Annexins, Enolase 1, Lamin A, and Glutathione-S-transferase omega 1. These results are consistent with recent studies indicating the crucial role of DEK in apoptosis pathway. We further demonstrated by ChIP analysis that knock-down of DEK caused hyperacetylation of histories around Prx VI promoter which is upregulated in our profile. Using immunoblotting analysis, we have demonstrated the modulation of other caspase-dependent apoptosis related proteins by DEK knock-down and further implicate its role in apoptosis pathway. J. Cell. Biochem. 106: 1048-1059, 2009. (c) 2009 Wiley-Liss, Inc.