An approach to further enhance the cellular productivity of exogenous protein hyper-producing Chinese hamster ovary (CHO) cells

Kiichiro Teruya, Yoshihito Daimon, Xiao Yan Dong, Yoshinori Katakura, Takumi Miura, Akira Ichikawa, Tsukasa Fujiki, Makiko Yamashita, Tetsuya Mori, Hideya Ohashi, Sanetaka Shirahata

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Abstract

The cell line D29, which was easily and rapidly established by the promoter-activated production and glutamine synthetase hybrid system, secreted recombinant human interleukin-6 (hIL-6) at a productivity rate of 39.5 μg 10-6 cells day-1, one of the highest reported levels worldwide. The productivity rate was about 130-fold higher than that of the cell line A7, which was established without both promoter activation and gene amplification. Although D29 cells had a high copy number and high mRNA level of the hIL-6 gene as well as a high secretion rate of hIL-6, large amounts of intracellular hIL-6 protein accumulated in D29 cells compared to A7 cells. Northern blotting analysis showed no change in the GRP78/BiP expression level in D29 cells. In contrast, an electrophoresis mobility shift assay revealed strong activation of NF-κB in D29 cells. These results suggest that large amounts of hIL-6 translated from large amounts of hIL-6 mRNA cause excess accumulation of intact hIL-6 in the endoplasmic reticulum (ER), and that subsequent negative feedback signals via the ER overload response inhibit hIL-6 protein secretion. To enhance the hIL-6 productivity rate of D29 cells by releasing the negative feedback signals, the effect of pyrrolidinedithiocarbamate, an inhibitor of NF-κB activation, was examined. Suppression of NF-κB activation in D29 cells produced a 25% augmentation of the hIL-6 productivity rate. Therefore, in highly productive cells like D29 cells, the release of negative feedback signals could increase the total amount of recombinant protein secretion.

Original languageEnglish
Pages (from-to)29-36
Number of pages8
JournalCytotechnology
Volume47
Issue number1-3
DOIs
Publication statusPublished - Jan 1 2005

All Science Journal Classification (ASJC) codes

  • Biotechnology
  • Bioengineering
  • Biomedical Engineering
  • Clinical Biochemistry
  • Cell Biology

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    Teruya, K., Daimon, Y., Dong, X. Y., Katakura, Y., Miura, T., Ichikawa, A., Fujiki, T., Yamashita, M., Mori, T., Ohashi, H., & Shirahata, S. (2005). An approach to further enhance the cellular productivity of exogenous protein hyper-producing Chinese hamster ovary (CHO) cells. Cytotechnology, 47(1-3), 29-36. https://doi.org/10.1007/s10616-005-3765-4