Reduced apoptosis in Chinese hamster ovary cells via optimized CRISPR interference

Kai Xiong, Kim Fabiano Marquart, Karen Julie la Cour Karottki, Shangzhong Li, Isaac Shamie, Jae Seong Lee, Signe Gerling, Nan Cher Yeo, Alejandro Chavez, Gyun Min Lee, Nathan E Lewis, Helene Faustrup Kildegaard*

*Corresponding author for this work

Research output: Contribution to journalJournal articleResearchpeer-review

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Chinese hamster ovary (CHO) cells are widely used for biopharmaceutical protein production. One challenge limiting CHO cell productivity is apoptosis stemming from cellular stress during protein production. Here we applied CRISPR interference (CRISPRi) to downregulate the endogenous expression of apoptotic genes Bak, Bax, and Casp3 in CHO cells. In addition to reduced apoptosis, mitochondrial membrane integrity was improved and the caspase activity was reduced. Moreover, we optimized the CRISPRi system to enhance the gene repression efficiency in CHO cells by testing different repressor fusion types. An improved Cas9 repressor has been identified by applying C-terminal fusion of a bipartite repressor domain, KRAB-MeCP2, to nuclease-deficient Cas9. These results collectively demonstrate that CHO cells can be rescued from cell apoptosis by targeted gene repression using the CRISPRi system.
Original languageEnglish
JournalBiotechnology and Bioengineering
Issue number7
Pages (from-to)1813-1819
Number of pages7
Publication statusPublished - 2019


  • Bak
  • Bax
  • CHO
  • Casp3
  • apoptosis
  • caspase
  • mitochondrial membrane integrity


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