Quantitative phosphoproteomics of CXCL12 (SDF-1) signaling.
Quantitative phosphoproteomics of CXCL12 (SDF-1) signaling.
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DOI：
10.1371/journal.pone.0024918
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发表时间：
2011
期刊：
影响因子：
3.7
通讯作者：
Doms RW
中科院分区：
文献类型：
--
作者：
Wojcechowskyj JA;Lee JY;Seeholzer SH;Doms RW
CXCL12 （SDF-1）是一种结合并通过7个跨膜受体CXCR4发出信号的趋化因子。CXCL12/CXCR4信号轴与癌症转移和人类免疫缺陷病毒1型（HIV-1）感染有关，对CXCL12/CXCR4信号通路的更全面了解可能有助于开发这些疾病的治疗方法。以质谱为基础的磷蛋白质组学已成为研究信号网络的重要工具。我们采用细胞培养氨基酸稳定同位素标记（SILAC）定量磷酸化蛋白质组学方法检测了人淋巴细胞CEM细胞系中CXCL12/CXCR4信号轴。我们从1,673个蛋白中定量了4,074个独特的SILAC对，89个磷酸化肽被认为在生物复制中对cxcl12有反应。我们的研究证实了几个已建立的cxcl12响应磷酸化位点，如AKT （pS473）和ERK2 （pY204）。我们还通过Western blot验证了两个新的cxcl12响应磷酸化位点，stathmin （pS16）和AKT1S1 （pT246）。通路分析和与其他磷酸化蛋白质组学数据集的比较显示，来自CXCL12响应磷酸化位点的基因丰富于细胞通路，如T细胞活化、表皮生长因子和哺乳动物雷帕霉素靶蛋白（mTOR）信号通路，这些通路先前被认为与CXCL12/CXCR4信号通路相关。我们研究的一些新的cxcl12应答磷酸化蛋白也与细胞迁移和HIV-1感染有关，因此为癌症转移和HIV-1治疗的发展提供了一个有吸引力的潜在靶点列表，并进一步加深了我们对趋化因子信号调节的理解。
CXCL12 (SDF-1) is a chemokine that binds to and signals through the seven transmembrane receptor CXCR4. The CXCL12/CXCR4 signaling axis has been implicated in both cancer metastases and human immunodeficiency virus type 1 (HIV-1) infection and a more complete understanding of CXCL12/CXCR4 signaling pathways may support efforts to develop therapeutics for these diseases. Mass spectrometry-based phosphoproteomics has emerged as an important tool in studying signaling networks in an unbiased fashion. We employed stable isotope labeling with amino acids in cell culture (SILAC) quantitative phosphoproteomics to examine the CXCL12/CXCR4 signaling axis in the human lymphoblastic CEM cell line. We quantified 4,074 unique SILAC pairs from 1,673 proteins and 89 phosphopeptides were deemed CXCL12-responsive in biological replicates. Several well established CXCL12-responsive phosphosites such as AKT (pS473) and ERK2 (pY204) were confirmed in our study. We also validated two novel CXCL12-responsive phosphosites, stathmin (pS16) and AKT1S1 (pT246) by Western blot. Pathway analysis and comparisons with other phosphoproteomic datasets revealed that genes from CXCL12-responsive phosphosites are enriched for cellular pathways such as T cell activation, epidermal growth factor and mammalian target of rapamycin (mTOR) signaling, pathways which have previously been linked to CXCL12/CXCR4 signaling. Several of the novel CXCL12-responsive phosphoproteins from our study have also been implicated with cellular migration and HIV-1 infection, thus providing an attractive list of potential targets for the development of cancer metastasis and HIV-1 therapeutics and for furthering our understanding of chemokine signaling regulation by reversible phosphorylation.