%0 Journal Article %1 Pieper2017 %A Pieper, Lisa A. %A Strotbek, Michaela %A Wenger, Till %A Gamer, Martin %A Olayioye, Monilola A. %A Hausser, Angelika %D 2017 %I Academic Press %J Metabolic Engineering %K 2017 hausser izi olayioye %P 69--79 %R 10.1016/j.ymben.2017.01.003 %T Secretory pathway optimization of CHO producer cells by co-engineering of the mitosRNA-1978 target genes CerS2 and Tbc1D20 %U https://www.sciencedirect.com/science/article/pii/S1096717616301744?via\%3Dihub %V 40 %X Chinese Hamster Ovary (CHO) cells are the most commonly used host for the production of biopharmaceuticals. Although transcription and translation engineering strategies have been employed to generate high-producer cell clones, the secretory pathway still remains a bottleneck in cellular productivity. In this study we show that ectopic expression of a human mitochondrial genome-encoded small RNA (mitosRNA-1978) in an IgG expressing CHO cell line strongly improved specific productivity by functioning in a microRNA-like fashion. By next generation sequencing we identified two endoplasmic reticulum (ER)-localized proteins, Ceramide Synthase 2 (CerS2) and the Rab1 GAP Tbc domain family member 20 (Tbc1D20), as target genes of mitosRNA-1978. Combined transient siRNA-mediated knockdown of CerS2 and Tbc1D20 resulted in increased specific productivity of CHO-IgG cells, thus recapitulating the mitosRNA-1978 phenotype. In support of a function in vesicular trafficking at the level of the ER, we provide evidence for altered cellular ceramide composition upon CerS2 knockdown and increased activity of Rab1 in CHO-IgG cells depleted of Tbc1D20. Importantly, in a fed-batch process, the combined stable knockdown of CerS2 and Tbc1D20 in CHO-IgG cells resulted in dramatically increased antibody production which was accompanied by enhanced cell growth. Thus, by identifying mitosRNA-1978 target genes in combination with an informed shRNA-mediated co-engineering approach we successfully optimized the secretory capacity of CHO producer cells used for the manufacturing of therapeutic proteins.

@article{Pieper2017, abstract = {Chinese Hamster Ovary (CHO) cells are the most commonly used host for the production of biopharmaceuticals. Although transcription and translation engineering strategies have been employed to generate high-producer cell clones, the secretory pathway still remains a bottleneck in cellular productivity. In this study we show that ectopic expression of a human mitochondrial genome-encoded small RNA (mitosRNA-1978) in an IgG expressing CHO cell line strongly improved specific productivity by functioning in a microRNA-like fashion. By next generation sequencing we identified two endoplasmic reticulum (ER)-localized proteins, Ceramide Synthase 2 (CerS2) and the Rab1 GAP Tbc domain family member 20 (Tbc1D20), as target genes of mitosRNA-1978. Combined transient siRNA-mediated knockdown of CerS2 and Tbc1D20 resulted in increased specific productivity of CHO-IgG cells, thus recapitulating the mitosRNA-1978 phenotype. In support of a function in vesicular trafficking at the level of the ER, we provide evidence for altered cellular ceramide composition upon CerS2 knockdown and increased activity of Rab1 in CHO-IgG cells depleted of Tbc1D20. Importantly, in a fed-batch process, the combined stable knockdown of CerS2 and Tbc1D20 in CHO-IgG cells resulted in dramatically increased antibody production which was accompanied by enhanced cell growth. Thus, by identifying mitosRNA-1978 target genes in combination with an informed shRNA-mediated co-engineering approach we successfully optimized the secretory capacity of CHO producer cells used for the manufacturing of therapeutic proteins.}, added-at = {2023-06-29T13:07:55.000+0200}, author = {Pieper, Lisa A. and Strotbek, Michaela and Wenger, Till and Gamer, Martin and Olayioye, Monilola A. and Hausser, Angelika}, biburl = {https://puma.ub.uni-stuttgart.de/bibtex/278dbbf1d08bf5bd5b0a62afedf7748df/fabian}, doi = {10.1016/j.ymben.2017.01.003}, interhash = {d9f23a9854882d494c2ece4a0dad20d9}, intrahash = {78dbbf1d08bf5bd5b0a62afedf7748df}, issn = {10967184}, journal = {Metabolic Engineering}, keywords = {2017 hausser izi olayioye}, month = mar, pages = {69--79}, pmid = {28088541}, publisher = {Academic Press}, timestamp = {2023-06-29T13:07:55.000+0200}, title = {{Secretory pathway optimization of CHO producer cells by co-engineering of the mitosRNA-1978 target genes CerS2 and Tbc1D20}}, url = {https://www.sciencedirect.com/science/article/pii/S1096717616301744?via{\%}3Dihub}, volume = 40, year = 2017 }