Epigenetic regulatory elements: Recent advances in understanding their mode of action and use for recombinant protein production in mammalian cells

Harraghy, Niamh; Calabrese, David; Fisch, Igor; Girod, Pierre‐Alain; LeFourn, Valérie; Regamey, Alexandre; Mermod, Nicolas

doi:10.1002/biot.201400649

Cited by 30 publications

(28 citation statements)

References 94 publications

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“…The source of MARs, the inserted position of MARs, and the type of host cells may affect transgene expression by MAR [5,30,37,38]. Moreover, gene copy numbers are related to transgene expression [39], and the methylation of DNA may reduce transgene expression [40][41][42]. Some reports have demonstrated that MARs improve the expression of transgenes through the recombination pathway of synthesis-dependent microhomology-mediated endjoining (MMEJ) [8,43,44].…”

Section: Discussionmentioning

confidence: 99%

A chimeric HS4 insulator–scaffold attachment region enhances transgene expression in transfected Chinese hamster ovary cells

et al. 2017

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Chinese hamster ovary ( CHO ) cells are one of the most commonly used expression systems for the production of recombinant proteins but low levels of transgene expression and transgene silencing are frequently encountered. Epigenetic regulatory elements such as the chicken β‐globin locus control region hypersensitive site 4 (HS4) and scaffold/matrix attachment regions (S/MARs) have positive effects on transgene expression. In this study, a chimeric HS 4‐ SAR was cloned upstream or downstream of an enhanced green fluorescent protein ( eGFP ) expression cassette in a eukaryotic vector, and the resulting vectors were transfected into CHO cells. eGFP was detected by flow cytometry. Real‐time quantitative PCR (qPCR) was used to determine copy numbers of the stably transfected cells. And fluorescence in situ hybridization ( FISH ) was used to detect the status of vector in the host cell chromosome. The results showed that HS 4‐ SAR positioned downstream of the expression cassette could enhance eGFP expression by 4.83‐fold compared with the control vector. There may not be a relationship between transgene copy number and gene expression level. HS 4‐ SAR did not appear to alter the integration of the transgene into the host cell chromosome or its position in the chromosome. We found a synthetic chimeric HS 4‐ SAR positively increased transgene expression in CHO cells.

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Section: Discussionmentioning

confidence: 99%

A chimeric HS4 insulator–scaffold attachment region enhances transgene expression in transfected Chinese hamster ovary cells

et al. 2017

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“…opening element (UCOE), or other insulator elements in the expression vector [24 ]. Strong constitutive promoters such as the mouse cytomegalovirus major immediate early promoter/enhancer (mCMV-MIE), the human elongation factor-1 alpha promoter (hEF-1a), and the SV40 early promoter/enhancer (SV40E) are recommended as alternatives to the hCMV-MIE [25,26].…”

Section: Expression Vector Elementsmentioning

confidence: 99%

Recombinant protein production from stable mammalian cell lines and pools

Hacker

Balasubramanian

2016

Current Opinion in Structural Biology

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“…In the following section, each element will be briefly discussed (see also Table ). For further details the reader is referred to the review …”

Section: Strategies To Improve Transgene Transcription: Chromatin‐modmentioning

confidence: 99%

Engineered and Natural Promoters and Chromatin‐Modifying Elements for Recombinant Protein Expression in CHO Cells

Romanova

Noll

2017

Biotechnology Journal

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Promoters play a pivotal role in integrating and processing the signals related to transcription initiation. Strong natural viral promoters, such as hCMV or SV40E, have been routinely employed to achieve a high rate of gene expression in ubiquitously used Chinese hamster ovary (CHO) cells. However, viral promoters are susceptible to epigenetic silencing and lack precise regulation levers. This has paved the way to more sensible control elements: endogenous, inducible, and synthetic promoters. In this review we summarize and discuss the use of natural viral, mammalian, and endogenous promoters, as well as recent advances in synthetic promoters and inducible systems for protein expression in CHO cells. Not only the level of transcription, but its long-term stability is crucial for recombinant protein production. Epigenetic chromatin-modifying elements, such as ubiquitously acting chromatin opening elements (UCOEs), matrix attachment regions (MARs), insulators and stabilizing anti-repressors (STARs) significantly improve transcription levels over extended cultivation time and are also discussed here. This review provides up-to date information to facilitate the choice of a suitable promoter and adjacent chromatin-modifying elements to maximize transgene expression as well as ensure long-term expression stability in CHO cell culture.

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Epigenetic regulatory elements: Recent advances in understanding their mode of action and use for recombinant protein production in mammalian cells

Cited by 30 publications

References 94 publications

A chimeric HS4 insulator–scaffold attachment region enhances transgene expression in transfected Chinese hamster ovary cells

A chimeric HS4 insulator–scaffold attachment region enhances transgene expression in transfected Chinese hamster ovary cells

Recombinant protein production from stable mammalian cell lines and pools

Engineered and Natural Promoters and Chromatin‐Modifying Elements for Recombinant Protein Expression in CHO Cells

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