Knockout of the caspase 8-associated protein 2 gene improves recombinant protein expression in HEK293 cells through up-regulation of the cyclin-dependent kinase inhibitor 2A gene.
Biotechnol Bioeng
; 118(1): 186-198, 2021 01.
Article
in En
| MEDLINE
| ID: mdl-32910455
ABSTRACT
Cell lines used in bioproduction are routinely engineered to improve their production efficiency. Numerous strategies, such as random mutagenesis, RNA interference screens, and transcriptome analyses have been employed to identify effective engineering targets. A genome-wide small interfering RNA screen previously identified the CASP8AP2 gene as a potential engineering target for improved expression of recombinant protein in the HEK293 cell line. Here, we validate the CASP8AP2 gene as an engineering target in HEK293 cells by knocking it out using CRISPR/Cas9 genome editing and assessing the effect of its knockout on recombinant protein expression, cell growth, cell viability, and overall gene expression. HEK293 cells lacking CASP8AP2 showed a seven-fold increase in specific expression of recombinant luciferase and a 2.5-fold increase in specific expression of recombinant SEAP, without significantly affecting cell growth and viability. Transcriptome analysis revealed that the deregulation of the cell cycle, specifically the upregulation of the cyclin-dependent kinase inhibitor 2A (CDKN2A) gene, contributed to the improvement in recombinant protein expression in CASP8AP2 deficient cells. The results validate the CASP8AP2 gene is a viable engineering target for improved recombinant protein expression in the HEK293 cell line.
Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Calcium-Binding Proteins
/
Gene Expression Regulation, Enzymologic
/
Up-Regulation
/
Cyclin-Dependent Kinase Inhibitor p16
/
Apoptosis Regulatory Proteins
Type of study:
Prognostic_studies
/
Risk_factors_studies
Limits:
Humans
Language:
En
Journal:
Biotechnol Bioeng
Year:
2021
Document type:
Article
Affiliation country: