Temporal profiling of the adipocyte proteome during differentiation using a five-plex SILAC based strategy.
Temporal profiling of the adipocyte proteome during differentiation using a five-plex SILAC based strategy.
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DOI：
10.1021/pr800650r
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发表时间：
2009-01
影响因子：
4.4
通讯作者：
Pandey, Akhilesh
中科院分区：
文献类型：
--
作者：
Molina, Henrik;Yang, Yi;Ruch, Travis;Kim, Jae-Woo;Mortensen, Peter;Otto, Tamara;Nalli, Anuradha;Tang, Qi-Qun;Lane, M. Daniel;Chaerkady, Raghothama;Pandey, Akhilesh
脂肪组织在人体内具有重要的分泌和内分泌功能。在过去的几年里，脂肪细胞分化的调控一直在积极地使用转录组学方法进行。定量蛋白质组学已成为一种很有前途的方法，以获得生物过程，如分化的时间配置文件。细胞培养中氨基酸稳定同位素标记法（SILAC）是一种简单、可靠的体内蛋白质标记方法。在这里，我们描述了一个五重SILAC实验的开发和应用，使用四种不同的重稳定同位素形式的精氨酸研究脂肪细胞分化过程中的核蛋白质组和分泌组。使用四极杆飞行时间仪器的串联质谱分析导致从这两个蛋白质组中鉴定出总共882种蛋白质。在这些蛋白质中，427种基于一种或多种允许定量的含精氨酸的肽鉴定。除了先前报道的在脂肪形成过程中差异表达的分子（例如脂联素和脂蛋白脂肪酶）之外，我们还鉴定了几种在脂肪细胞分化过程中差异表达的蛋白质，这些蛋白质先前没有被记录。例如，THO复合物4（T细胞受体α增强子复合物中的背景依赖性转录激活因子）在脂肪形成的中期显示最高表达，而SNF2 α（染色质重塑蛋白）在脂肪形成开始时下调，并在随后的时间点保持如此。本研究使用5重SILAC来研究动力学，说明了这种方法以时间方式鉴定差异表达蛋白的能力。
The adipose tissue has important secretory and endocrine functions in humans. The regulation of adipocyte differentiation has been actively pursued using transcriptomic methods over the last several years. Quantitative proteomics has emerged as a promising approach to obtain temporal profiles of biological processes such as differentiation. Stable isotope labeling with amino acids in cell culture (SILAC) is a simple and robust method for labeling proteins in vivo. Here, we describe the development and application of a five-plex SILAC experiment using four different heavy stable isotopic forms of arginine to study the nuclear proteome and the secretome during the course of adipocyte differentiation. Tandem mass spectrometry analysis using a quadrupole time-of-flight instrument resulted in identification of a total 882 proteins from these two proteomes. Of these proteins, 427 were identified on the basis of one or more arginine containing peptides that allowed quantitation. In addition to previously reported molecules that are differentially expressed during the process of adipogenesis (e.g. adiponectin and lipoprotein lipase), we identified several proteins whose differential expression during adipocyte differentiation has not been documented previously. For example, THO complex 4, a context-dependent transcriptional activator in the T-cell receptor alpha enhancer complex, showed highest expression at middle stage of adipogenesis while SNF2 alpha, a chromatin remodeling protein, was downregulated upon initiation of adipogenesis and remained so during subsequent time points. This study using a 5-plex SILAC to investigate dynamics illustrates the power of this approach to identify differentially expressed proteins in a temporal fashion.
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影响因子：
3.5
作者：
HUA, XX;ENERBACK, S;GIMBLE, JM
通讯作者：
GIMBLE, JM
影响因子：
5.3
作者：
Salma, N;Xiao, HG;Imbalzano, AN
通讯作者：
Imbalzano, AN
影响因子：
10.5
作者：
Bruhn, L;Munnerlyn, A;Grosschedl, R
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Grosschedl, R
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作者：
Hummasti, Sarah;Tontonoz, Peter
通讯作者：
Tontonoz, Peter
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46.9
作者：
Blagoev, B;Ong, SE;Mann, M
通讯作者：
Mann, M