The Genomic Sequence of the Chinese Hamster Ovary (CHO) K1 cell line

BGI-Shenzhen, Shenzhen, People's Republic of China.
Nature Biotechnology (Impact Factor: 41.51). 07/2011; 29(8):735-41. DOI: 10.1038/nbt.1932
Source: PubMed


Chinese hamster ovary (CHO)-derived cell lines are the preferred host cells for the production of therapeutic proteins. Here we present a draft genomic sequence of the CHO-K1 ancestral cell line. The assembly comprises 2.45 Gb of genomic sequence, with 24,383 predicted genes. We associate most of the assembled scaffolds with 21 chromosomes isolated by microfluidics to identify chromosomal locations of genes. Furthermore, we investigate genes involved in glycosylation, which affect therapeutic protein quality, and viral susceptibility genes, which are relevant to cell engineering and regulatory concerns. Homologs of most human glycosylation-associated genes are present in the CHO-K1 genome, although 141 of these homologs are not expressed under exponential growth conditions. Many important viral entry genes are also present in the genome but not expressed, which may explain the unusual viral resistance property of CHO cell lines. We discuss how the availability of this genome sequence may facilitate genome-scale science for the optimization of biopharmaceutical protein production.

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Available from: Mikael R Andersen, Oct 08, 2015
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    • "2244.1) were the three most abundant signals. In contrast to native IgG, CHO cells do not produce the bisecting GlcNAc branch (which can be found on more than 10% of human IgG glycoforms [26] [27]) due to the lack of GlcNAc transferase III activity [28] [29]. On the other hand, recombinant IgGs from CHO cells show ranges of highmannose-type N-glycans from 1% to greater than 20% [30] "
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    ABSTRACT: Glycosylation is the most complex posttranslational modification. Thus, it contributes to versatile chemical compositions of proteins, leading to high amounts of protein species. The structural heterogeneity of glycoproteins was also described by the definition of glycoforms. We therefore introduced a new term called "glycoprotein species" to join the two concepts from different fields of biology. In this study, we further determined the theoretical numbers of glycoprotein species of two recombinant glycoproteins - a therapeutical antibody and the human protease inhibitor alpha-1-antitrypsin (A1AT) - based on structural analysis of their N-glycans. Moreover, we showed that variations in the used cell lines and their cultivation conditions strongly influence the number of glycoprotein species in case of recombinant A1AT production. Protein glycosylation is a major source for the huge amount of protein species. This study extends the sight of protein species by the following contributions: 1) The new term "glycoprotein species" was defined to introduce the concept of glycoforms into the field. 2) An estimation of the number of potential glycoprotein species of two particular glycoproteins was given. 3) The influence of production conditions for recombinant glycoproteins on glycoprotein species generation was displayed. Copyright © 2015. Published by Elsevier B.V.
    Journal of proteomics 08/2015; DOI:10.1016/j.jprot.2015.08.011 · 3.89 Impact Factor
    • "N-glycans on native human proteins typically include both terminal a-2,3 and a-2.6 sialic acid, whereas the majority of oligosaccharides present on recombinant proteins expressed in CHO cells exclusively display a-2,3 sialylation (Lee et al., 1989; Takeuchi et al., 1988). This difference is due to the lack of significant expression of the a-2,6 sialyltransferase gene in CHO cell lines (Lewis et al., 2013; Svensson et al., 1990; Xu et al., 2011). These differences can be important because a number of studies have demonstrated the importance of both sialic acid content and branching to the biological activity and in vivo circulatory activity of recombinant glycoproteins (Fukuda et al., 1989; Misaizu et al., 1995; Richards et al., 2010; Wide et al., 2009). "
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    ABSTRACT: Sialic acid, a terminal residue on complex N-glycans, and branching or antennarity can play key roles in both the biological activity and circulatory lifetime of recombinant glycoproteins of therapeutic interest. In order to examine the impact of glycosyltransferase expression on the N-glycosylation of recombinant erythropoietin (rEPO), a human α2,6-sialyltransferase (ST6Gal1) was expressed in Chinese hamster ovary (CHO-K1) cells. Sialylation increased on both EPO and CHO cellular proteins as observed by SNA lectin analysis, and HPLC profiling revealed that the sialic acid content of total glycans on EPO increased by 26%. The increase in sialic acid content was further verified by detailed profiling of the N-glycan structures using mass spectra (MS) analysis. In order to enhance antennarity/branching, UDP-N-acetylglucosamine: α-1,3-D-mannoside β1,4-N-acetylglucosaminyltransferase (GnTIV/Mgat4) and UDP-N-acetylglucosamine:α-1,6-D-mannoside β1,6-N-acetylglucosaminyltransferase (GnTV/Mgat5), was incorporated into CHO-K1 together with ST6Gal1. Tri- and tetraantennary N-glycans represented approximately 92% of the total N-glycans on the resulting EPO as measured using MS analysis. Furthermore, sialic acid content of rEPO from these engineered cells was increased ∼45% higher with tetra-sialylation accounting for ∼10% of total sugar chains compared to ∼3% for the wild-type parental CHO-K1. In this way, coordinated overexpression of these three glycosyltransferases for the first time in model CHO-K1 cell lines provides a mean for enhancing both N-glycan branching complexity and sialylation with opportunities to generate tailored complex N-glycan structures on therapeutic glycoproteins in the future. Biotechnol. Bioeng. 2015;9999: 1-8. © 2015 Wiley Periodicals, Inc. © 2015 Wiley Periodicals, Inc.
    Biotechnology and Bioengineering 07/2015; DOI:10.1002/bit.25650 · 4.13 Impact Factor
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    • "Genome references and statistics used for the identification of evolutionary conserved microRNAs. Lewis et al. (2013) Xu et al. (2011) Brinkrolf et al. (2013) K1-BB "
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    ABSTRACT: MicroRNAs are small non-coding RNAs that play a critical role in post-transcriptional control of gene expression. Recent publications of genomic sequencing data from the Chinese Hamster (CGR) and Chinese hamster ovary (CHO) cells provide new tools for the discovery of novel miRNAs in this important production system. Version 20 of the miRNA registry miRBase contains 307 mature miRNAs and 200 precursor sequences for CGR/CHO. We searched for evolutionary conserved miRNAs from miRBase v20 in recently published genomic data, derived from Chinese hamster and CHO cells, to further extend the list of known miRNAs. With our approach we could identify several hundred miRNA sequences in the genome. For several of these, the expression in CHO cells could be verified from multiple next-generation sequencing experiments. In addition, several hundred unexpressed miRNAs are awaiting further confirmation by testing for their transcription in different Chinese hamster tissues. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.
    Biotechnology and Bioengineering 02/2015; 112(7). DOI:10.1002/bit.25539 · 4.13 Impact Factor
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