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

研究成果: Contribution to journalArticle査読

4 被引用数 (Scopus)

抄録

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.

本文言語英語
ページ(範囲)29-36
ページ数8
ジャーナルCytotechnology
47
1-3
DOI
出版ステータス出版済み - 1 2005

All Science Journal Classification (ASJC) codes

  • バイオテクノロジー
  • バイオエンジニアリング
  • 生体医工学
  • 臨床生化学
  • 細胞生物学

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