Anti-cell death engineering of CHO cells: Co-overexpression of Bcl-2 for apoptosis inhibition, Beclin-1 for autophagy induction
Article first published online: 16 MAR 2013
Copyright © 2013 Wiley Periodicals, Inc.
Biotechnology and Bioengineering
Volume 110, Issue 8, pages 2195–2207, August 2013
How to Cite
Lee, J. S., Ha, T. K., Park, J. H. and Lee, G. M. (2013), Anti-cell death engineering of CHO cells: Co-overexpression of Bcl-2 for apoptosis inhibition, Beclin-1 for autophagy induction. Biotechnol. Bioeng., 110: 2195–2207. doi: 10.1002/bit.24879
- Issue published online: 25 JUN 2013
- Article first published online: 16 MAR 2013
- Accepted manuscript online: 22 FEB 2013 03:19PM EST
- Manuscript Accepted: 15 FEB 2013
- Manuscript Revised: 14 JAN 2013
- Manuscript Received: 8 AUG 2012
- Ministry of Education, Science, and Technology (MEST). Grant Numbers: R31-2008-000-10071-0, 2009-0082276, 2011-0031962
Genetic engineering approaches to inhibit cell death in Chinese hamster ovary (CHO) cell cultures have been limited primarily to anti-apoptosis engineering. Recently, autophagy has received attention as a new anti-cell death engineering target in addition to apoptosis. In order to achieve a more efficient protection of cells from the stressful culture conditions, the simultaneous targeting of anti-apoptosis and pro-autophagy in CHO cells (DG44) was attempted by co-overexpressing an anti-apoptotic protein, Bcl-2, and a key regulator of autophagy pathway, Beclin-1, respectively. Co-overexpression of Bcl-2 and Beclin-1 exhibited a longer culture period as well as higher viability during serum-free suspension culture, compared with the control (without co-overexpression of Bcl-2 and Beclin-1) and Bcl-2 overexpression only. In addition to the efficient inhibition of apoptosis by Bcl-2 overexpression, Beclin-1 overexpression successfully induced the increase in the autophagic marker protein, LC3-II, and autophagosome formation with the decrease in mTOR activity. Co-immunoprecipitation and qRT-PCR experiments revealed that the enforced expression of Beclin-1 increased Ulk1 expression and level of free-Beclin-1 that did not bind to the Bcl-2 despite the Bcl-2 overexpression. Under other stressful culture conditions such as treatment with sodium butyrate and hyperosmolality, co-overexpression of Bcl-2 and Beclin-1 also protected the cells from cell death more efficiently than Bcl-2 overexpression only, implying the potential of autophagy induction. Taken together, the data obtained here provide the evidence that pro-autophagy engineering together with anti-apoptosis engineering yields a synergistic effect and successfully enhances the anti-cell death engineering of CHO cells. Biotechnol. Bioeng. 2013; 110: 2195–2207. © 2013 Wiley Periodicals, Inc.