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

Research output: Contribution to journalArticlepeer-review

Abstract

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.

Original languageEnglish
Article number110936
Pages (from-to)110936
JournalCell Reports
Volume39
Issue number11
DOIs
Publication statusPublished - 14 Jun 2022

Bibliographical note

Funding Information:
This work was supported by the Knut and Alice Wallenberg Foundation ; AstraZeneca ; Swedish Foundation for Strategic Research (SSF); Swedish innovation agency Vinnova through AAVNova , CellNova , and AdBIOPRO ; and the Novo Nordisk Foundation (grant no. NNF10CC1016517 ). The computations/data handling was enabled by resources provided by the Swedish National Infrastructure for Computing (SNIC), partially funded by the Swedish Research Council through grant agreement no. 2018-05973 .

Funding Information:
This work was supported by the Knut and Alice Wallenberg Foundation; AstraZeneca; Swedish Foundation for Strategic Research (SSF); Swedish innovation agency Vinnova through AAVNova, CellNova, and AdBIOPRO; and the Novo Nordisk Foundation (grant no. NNF10CC1016517). The computations/data handling was enabled by resources provided by the Swedish National Infrastructure for Computing (SNIC), partially funded by the Swedish Research Council through grant agreement no. 2018-05973. Conceptualization, J.N. J.L.R. and J.R.; methodology, R.S. J.Z. N.M. M.M. J.L.R. and J.R.; formal analysis, R.S. J.Z. and M.M.; investigation, R.S. M.M. J.Z. V.C. R.R. N.M. P.-O.B. N.W. H.T. A.P. A.H. F.E. J.L.R. and J.R.; writing – original draft, R.S. J.Z. M.M. J.L.R. and J.R.; writing – review & editing, R.S. J.Z. M.M. N.M. P.-O.B. D.H. J.N. L.G. V.C. J.L.R. and J.R.; visualization, R.S. J.Z. L.G. and J.L.R.; supervision, D.H. T.S. J.N. J.L.R. and J.R. funding acquisition, J.N. and J.R. P.-O.B. is cofounder and CEO of Biocrine AB. L.G. and D.H. are employees of AstraZeneca and may own AstraZeneca stock or stock options.

Publisher Copyright:
© 2022 The Authors

Keywords

  • ATF6B
  • CP: Cell biology
  • CP: Molecular biology
  • Cell engineering
  • GFP
  • HEK293
  • erythropoietin
  • protein production
  • ribosome heterogeneity
  • secretory pathways

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