Large scale gene expression profiling of metabolic shift of mammalian cells in culture
Large scale gene expression profiling of metabolic shift of mammalian cells in culture
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DOI：
10.1016/j.jbiotec.2003.09.007
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发表时间：
2004-01-08
影响因子：
4.1
通讯作者：
Hu, WS
中科院分区：
文献类型：
--
作者：
Korke, R;Gatti, MD;Hu, WS
杂交瘤细胞在连续培养中的代谢状态随着培养起始的培养条件而变化。在代谢转变状态下，与分批培养或连续培养中没有代谢转变的细胞相比，细胞具有显著降低的葡萄糖和其他营养物消耗以及乳酸盐产生。本研究采用基因组学和蛋白质组学相结合的方法，探讨了代谢转变的分子机制。研究了来自连续培养物的细胞在两种不同的稳态下的葡萄糖消耗与乳酸盐产生的摩尔比（DeltaL/DeltaG）为0.08和1.4。采用AffysseGeneChips和cDNA微阵列技术鉴定差异表达的mRNA转录本，采用2D凝胶电泳-质谱法鉴定差异表达的蛋白质。代谢转变时葡萄糖代谢的降低伴随着参与其代谢的许多基因的基因表达的降低。然而，差异表达的基因的数量和代谢转变后的差异表达的程度相对适中。通过真实的时间PCR证实了代谢基因在转录水平上的表达变化。结果表明，代谢转变是反应水平的生化事件和转录和翻译水平的基因表达的综合效应。这种整合转录谱分析、蛋白质组学技术和生化分析的方法提供了对培养中哺乳动物细胞代谢的更全面的看法。(C)2003 Elsevier B. V.保留所有权利。
The metabolic state of hybridoma cells in continuous culture varies with the cultivation condition from which the culture is initiated. At a metabolically shifted state, cells have markedly reduced glucose and other nutrient consumption and lactate production as compared to cells in batch culture or in continuous culture without a metabolic shift. Taking a combined genomics and proteornics approach, we investigated the molecular mechanism of metabolic shift. Cells from continuous cultures at two different steady states with a glucose consumption to lactate production molar ratio (DeltaL/DeltaG) of 0.08 and 1.4 were studied. Affymetrix GeneChips as well as cDNA microarrays were employed to identify differentially expressed mRNA transcripts, and the differentially expressed proteins were identified using the 2D gel electrophoresis-mass spectrometry approach. The decrease in glucose metabolism upon metabolic shift is accompanied by a decrease in gene expression of a number of genes involved in its metabolism. However, the number of genes differentially expressed and the extent of differential expression upon metabolic shift are relatively moderate. The change in the expression of metabolic genes at the transcriptional level was confirmed by real time PCR. The results suggest that metabolic shift is a combined effect of both biochemical events at reaction level and gene expression at transcription and translation level. This approach of integrating transcriptional profiling, proteomic techniques and biochemical analysis provides a more global view of the metabolism of mammalian cells in culture. (C) 2003 Elsevier B.V. All rights reserved.