Toward stable gene expression in CHO cells Preventing promoter silencing with core CpG island elements
Maintaining high gene expression level during long-term culture is critical when producing therapeutic recombinant proteins using mammalian cells. Transcriptional silencing of promoters, most likely due to epigenetic events such as DNA methylation and histone modifications, is one of the major mecha...
| Main Authors: | , , , , |
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| Other Authors: | |
| Format: | Journal Article |
| Language: | English |
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2016
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| Online Access: | https://hdl.handle.net/10356/81451 http://hdl.handle.net/10220/40784 |
| _version_ | 1826118555416920064 |
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| author | Mariati Koh, Esther YC Yeo, Jessna HM Ho, Steven CL Yang, Yuansheng |
| author2 | School of Chemical and Biomedical Engineering |
| author_facet | School of Chemical and Biomedical Engineering Mariati Koh, Esther YC Yeo, Jessna HM Ho, Steven CL Yang, Yuansheng |
| author_sort | Mariati |
| collection | NTU |
| description | Maintaining high gene expression level during long-term culture is critical when producing therapeutic recombinant proteins using mammalian cells. Transcriptional silencing of promoters, most likely due to epigenetic events such as DNA methylation and histone modifications, is one of the major mechanisms causing production instability. Previous studies demonstrated that the core CpG island element (IE) from the hamster adenine phosphoribosyltransferase gene is effective to prevent DNA methylation. We generated one set of modified human cytomegalovirus (hCMV) promoters by insertion of one or two copies of IE in either forward or reverse orientations into different locations of the hCMV promoter. The modified hCMV with one copy of IE inserted between the hCMV enhancer and core promoter in reverse orientation (MR1) was most effective at enhancing expression stability in CHO cells without comprising expression level when compared with the wild type hCMV. We also found that insertion of IE into a chimeric murine CMV (mCMV) enhancer and human elongation factor-1α core (hEF) promoter in reverse orientation did not enhance expression stability, indicating that the effect of IE on expression stability is possibly promoter specific. |
| first_indexed | 2024-10-01T04:45:43Z |
| format | Journal Article |
| id | ntu-10356/81451 |
| institution | Nanyang Technological University |
| language | English |
| last_indexed | 2024-10-01T04:45:43Z |
| publishDate | 2016 |
| record_format | dspace |
| spelling | ntu-10356/814512023-12-29T06:48:13Z Toward stable gene expression in CHO cells Preventing promoter silencing with core CpG island elements Mariati Koh, Esther YC Yeo, Jessna HM Ho, Steven CL Yang, Yuansheng School of Chemical and Biomedical Engineering CHO core CpG island element gene silencing expression stability Maintaining high gene expression level during long-term culture is critical when producing therapeutic recombinant proteins using mammalian cells. Transcriptional silencing of promoters, most likely due to epigenetic events such as DNA methylation and histone modifications, is one of the major mechanisms causing production instability. Previous studies demonstrated that the core CpG island element (IE) from the hamster adenine phosphoribosyltransferase gene is effective to prevent DNA methylation. We generated one set of modified human cytomegalovirus (hCMV) promoters by insertion of one or two copies of IE in either forward or reverse orientations into different locations of the hCMV promoter. The modified hCMV with one copy of IE inserted between the hCMV enhancer and core promoter in reverse orientation (MR1) was most effective at enhancing expression stability in CHO cells without comprising expression level when compared with the wild type hCMV. We also found that insertion of IE into a chimeric murine CMV (mCMV) enhancer and human elongation factor-1α core (hEF) promoter in reverse orientation did not enhance expression stability, indicating that the effect of IE on expression stability is possibly promoter specific. ASTAR (Agency for Sci., Tech. and Research, S’pore) Accepted version 2016-06-23T08:36:52Z 2019-12-06T14:31:17Z 2016-06-23T08:36:52Z 2019-12-06T14:31:17Z 2014 Journal Article Mariati, Koh, E. Y., Yeo, J. H., Ho, S. C., & Yang, Y. (2014). Toward stable gene expression in CHO cells. Bioengineered, 5(5), 340-345. 1949-1018 https://hdl.handle.net/10356/81451 http://hdl.handle.net/10220/40784 10.4161/bioe.32111 25482237 en Bioengineered © 2014 Landes Bioscience. This is the author created version of a work that has been peer reviewed and accepted for publication in Bioengineered, published by Taylor & Francis on behalf of Landes Bioscience. It incorporates referee’s comments but changes resulting from the publishing process, such as copyediting, structural formatting, may not be reflected in this document. The published version is available at: [http://dx.doi.org/10.4161/bioe.32111]. 6 p. application/pdf |
| spellingShingle | CHO core CpG island element gene silencing expression stability Mariati Koh, Esther YC Yeo, Jessna HM Ho, Steven CL Yang, Yuansheng Toward stable gene expression in CHO cells Preventing promoter silencing with core CpG island elements |
| title | Toward stable gene expression in CHO cells Preventing promoter silencing with core CpG island elements |
| title_full | Toward stable gene expression in CHO cells Preventing promoter silencing with core CpG island elements |
| title_fullStr | Toward stable gene expression in CHO cells Preventing promoter silencing with core CpG island elements |
| title_full_unstemmed | Toward stable gene expression in CHO cells Preventing promoter silencing with core CpG island elements |
| title_short | Toward stable gene expression in CHO cells Preventing promoter silencing with core CpG island elements |
| title_sort | toward stable gene expression in cho cells preventing promoter silencing with core cpg island elements |
| topic | CHO core CpG island element gene silencing expression stability |
| url | https://hdl.handle.net/10356/81451 http://hdl.handle.net/10220/40784 |
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