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  • Ribosome profiling-guided depletion of an mRNA increases cell growth rate and protein secretion.

Ribosome profiling-guided depletion of an mRNA increases cell growth rate and protein secretion.

Scientific reports (2017-01-17)
Thomas Beuchert Kallehauge, Shangzhong Li, Lasse Ebdrup Pedersen, Tae Kwang Ha, Daniel Ley, Mikael Rørdam Andersen, Helene Faustrup Kildegaard, Gyun Min Lee, Nathan E Lewis
ABSTRACT

Recombinant protein production coopts the host cell machinery to provide high protein yields of industrial enzymes or biotherapeutics. However, since protein translation is energetically expensive and tightly controlled, it is unclear if highly expressed recombinant genes are translated as efficiently as host genes. Furthermore, it is unclear how the high expression impacts global translation. Here, we present the first genome-wide view of protein translation in an IgG-producing CHO cell line, measured with ribosome profiling. Through this we found that our recombinant mRNAs were translated as efficiently as the host cell transcriptome, and sequestered up to 15% of the total ribosome occupancy. During cell culture, changes in recombinant mRNA translation were consistent with changes in transcription, demonstrating that transcript levels influence specific productivity. Using this information, we identified the unnecessary resistance marker NeoR to be a highly transcribed and translated gene. Through siRNA knock-down of NeoR, we improved the production- and growth capacity of the host cell. Thus, ribosomal profiling provides valuable insights into translation in CHO cells and can guide efforts to enhance protein production.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Monoclonal Anti-Vinculin antibody produced in mouse, clone hVIN-1, ascites fluid
Sigma-Aldrich
TWEEN® 20, viscous liquid
Sigma-Aldrich
ECO TWEEN® 20, viscous liquid