Enhanced metabolism and negative regulation of ER stress support higher erythropoietin production in HEK293 cells
Rasool Saghaleyni,
Magdalena Malm,
Noah Moruzzi,
Jan Zrimec,
Ronia Razavi,
Num Wistbacka,
Hannes Thorell,
Anton Pintar,
Andreas Hober,
Fredrik Edfors,
Veronique Chotteau,
Per-Olof Berggren,
Luigi Grassi,
Aleksej Zelezniak,
Thomas Svensson,
Diane Hatton,
Jens Nielsen,
Jonathan L. Robinson,
Johan Rockberg
Affiliations
Rasool Saghaleyni
Department of Biology and Biological Engineering, Chalmers University of Technology, 412 96 Gothenburg, Sweden
Magdalena Malm
KTH - Royal Institute of Technology, School of Engineering Sciences in Chemistry, Biotechnology, and Health, Department of Protein Science, 106 91 Stockholm, Sweden
Noah Moruzzi
The Rolf Luft Research Center for Diabetes and Endocrinology, Department of Molecular Medicine and Surgery, Karolinska Institute, 17176 Stockholm, Sweden
Jan Zrimec
Department of Biology and Biological Engineering, Chalmers University of Technology, 412 96 Gothenburg, Sweden
Ronia Razavi
KTH - Royal Institute of Technology, School of Engineering Sciences in Chemistry, Biotechnology, and Health, Department of Protein Science, 106 91 Stockholm, Sweden
Num Wistbacka
KTH - Royal Institute of Technology, School of Engineering Sciences in Chemistry, Biotechnology, and Health, Department of Protein Science, 106 91 Stockholm, Sweden
Hannes Thorell
KTH - Royal Institute of Technology, School of Engineering Sciences in Chemistry, Biotechnology, and Health, Department of Protein Science, 106 91 Stockholm, Sweden
Anton Pintar
KTH - Royal Institute of Technology, School of Engineering Sciences in Chemistry, Biotechnology, and Health, Department of Protein Science, 106 91 Stockholm, Sweden
Andreas Hober
Science for Life Laboratory, KTH - Royal Institute of Technology, 171 65 Solna, Sweden
Fredrik Edfors
Science for Life Laboratory, KTH - Royal Institute of Technology, 171 65 Solna, Sweden
Veronique Chotteau
KTH - Royal Institute of Technology, School of Engineering Sciences in Chemistry, Biotechnology, and Health, Department of Industrial Biotechnology, 106 91 Stockholm, Sweden
Per-Olof Berggren
The Rolf Luft Research Center for Diabetes and Endocrinology, Department of Molecular Medicine and Surgery, Karolinska Institute, 17176 Stockholm, Sweden
Luigi Grassi
Cell Culture & Fermentation Sciences, BioPharmaceutical Development, BioPharmaceuticals R&D, AstraZeneca, Cambridge, UK
Aleksej Zelezniak
Department of Biology and Biological Engineering, Chalmers University of Technology, 412 96 Gothenburg, Sweden
Thomas Svensson
Department of Biology and Biological Engineering, Chalmers University of Technology, 412 96 Gothenburg, Sweden; Department of Biology and Biological Engineering, National Bioinformatics Infrastructure Sweden, Science for Life Laboratory, Chalmers University of Technology, Kemivägen 10, 41258 Gothenburg, Sweden
Diane Hatton
Cell Culture & Fermentation Sciences, BioPharmaceutical Development, BioPharmaceuticals R&D, AstraZeneca, Cambridge, UK
Jens Nielsen
Department of Biology and Biological Engineering, Chalmers University of Technology, 412 96 Gothenburg, Sweden; Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, 2800 Kongens Lyngby, Denmark
Jonathan L. Robinson
Department of Biology and Biological Engineering, Chalmers University of Technology, 412 96 Gothenburg, Sweden; Department of Biology and Biological Engineering, National Bioinformatics Infrastructure Sweden, Science for Life Laboratory, Chalmers University of Technology, Kemivägen 10, 41258 Gothenburg, Sweden; Corresponding author
Johan Rockberg
KTH - Royal Institute of Technology, School of Engineering Sciences in Chemistry, Biotechnology, and Health, Department of Protein Science, 106 91 Stockholm, Sweden; Corresponding author
Summary: Recombinant protein production can cause severe stress on cellular metabolism, resulting in limited titer and product quality. To investigate cellular and metabolic characteristics associated with these limitations, we compare HEK293 clones producing either erythropoietin (EPO) (secretory) or GFP (non-secretory) protein at different rates. Transcriptomic and functional analyses indicate significantly higher metabolism and oxidative phosphorylation in EPO producers compared with parental and GFP cells. In addition, ribosomal genes exhibit specific expression patterns depending on the recombinant protein and the production rate. In a clone displaying a dramatically increased EPO secretion, we detect higher gene expression related to negative regulation of endoplasmic reticulum (ER) stress, including upregulation of ATF6B, which aids EPO production in a subset of clones by overexpression or small interfering RNA (siRNA) knockdown. Our results offer potential target pathways and genes for further development of the secretory power in mammalian cell factories.
