Microarray and Proteomics Expression Profiling Identifies Several Candidates, Including the Valosin-Containing Protein (VCP), Involved in Regulating High Cellular Growth Rate in Production CHO Cell Lines
Microarray and Proteomics Expression Profiling Identifies Several Candidates, Including the Valosin-Containing Protein (VCP), Involved in Regulating High Cellular Growth Rate in Production CHO Cell Lines
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DOI：
10.1002/bit.22670
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发表时间：
2010-05-01
影响因子：
3.8
通讯作者：
Clynes, Martin
中科院分区：
文献类型：
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作者：
Doolan, Padraig;Meleady, Paula;Clynes, Martin
导致活生物质快速积累的高细胞生长速率（pc）是在规模扩大操作和生产培养的早期阶段期间的期望表型。为了鉴定在中国仓鼠卵巢（CHO）细胞中有助于更高生长速率的基因和蛋白质，使用微阵列和蛋白质组表达谱分析的组合方法在显示快速或缓慢生长速率的两对匹配的CHO生产细胞系上进行。微阵列和蛋白质组数据的统计分析分别导致118个基因转录本和58个蛋白质的鉴定，这些基因转录本和蛋白质在快速生长细胞和缓慢生长细胞之间差异表达。两个数据集的重叠比较确定了与CHO中高生长速率表型相关的21个候选物的优先级列表。对这些候选物中的五个的功能分析（通过siRNA）鉴定了含缬氨肽的蛋白质（VCP）对CHO细胞生长和活力具有实质性影响。HSPB 1和ENO 1的敲低也对细胞生长有影响（分别为阴性和阳性）。使用基因敲低（siRNA）和过表达（cDNA）在CHO中进行的进一步功能验证证实，改变的VCP表达影响CHO细胞增殖，表明VCP和此处鉴定的其他基因和蛋白质可能在对数工程策略期间在CHO细胞生长的调节中发挥重要作用。Biotechnol. Bioeng. 2010;106：42-56. (C)2010 Wiley Periodicals，Inc.
A high rate of cell growth (pc) leading to rapid accumulation of viable biomass is a desirable phenotype during scale up operations and the early stages of production cultures. In order to identify genes and proteins that contribute to higher growth rates in Chinese hamster ovary (CHO) cells, a combined approach using microarray and proteomic expression profiling analysis was carried out on two matched pairs of CHO production cell lines that displayed either fast or slow growth rates. Statistical analysis of the microarray and proteomic data separately resulted in the identification of 118 gene transcripts and 58 proteins that were differentially expressed between the fast- and slow-growing cells. Overlap comparison of both datasets identified a priority list of 21 candidates associated with a high growth rate phenotype in CHO. Functional analysis (by siRNA) of five of these candidates identified the valosin-containing protein (VCP) as having a substantial impact on CHO cell growth and viability. Knockdown of HSPB1 and ENO1 also had an effect on cell growth (negative and positive, respectively). Further functional validation in CHO using both gene knockdown (siRNA) and overexpression (cDNA) confirmed that altered VCP expression impacted CHO cell proliferation, indicating that VCP and other genes and proteins identified here may play an important role in the regulation of CHO cell growth during log engineering strategies. Biotechnol. Bioeng. 2010;106: 42-56. (C) 2010 Wiley Periodicals, Inc.