title: Temperature-sensitive miR-483 is a conserved regulator of recombinant protein and viral vector production in mammalian cells.
abstract: Cell engineering and bioprocess optimizations such as low temperature cultivation represent powerful tools to improve cellular performance and product yields of mammalian production cells. Besides monoclonal antibodies (mABs), novel biotherapeutic formats such as viral vectors will gain increasing importance. Here, we demonstrate that similar to Chinese hamster ovary (CHO) cells, product yields of recombinant adeno-associated virus (rAAV) producing HeLa cells can be markedly increased by low temperature cultivation. MicroRNAs (miRNAs) are small non-coding RNAs that critically regulate cell phenotypes. We thus investigated differential miRNA expression in response to mild hypothermia in CHO and HeLa production cells. We discovered miR-483 to be substantially up-regulated upon temperature down-shift in both cell types. Functional validation experiments revealed that introduction of miR-483 mimics led to a significant increase in both rAAV and mAB production in HeLa and CHO cells, respectively. Furthermore, inhibition of miR-483 up-regulation during mild hypothermia significantly decreased product yields, suggesting that miR-483 is a key regulator of cellular productivity in mammalian cells. In addition, miRNA target gene identification indicated that miR-483 might regulate genes directly involved in cellular survival and protein expression. Our results highlight that miR-483 is a valuable tool for product-independent engineering of mammalian production cells.
authors.affiliation: Department of Gene Therapy, Ulm University, Ulm, Germany.|Rentschler Biotechnologie GmbH, Erwin-Rentschler-Str. 21, Laupheim, Germany.
authors.forename: Verena V
authors.initials: VV
authors.lastname: Emmerling
authors.affiliation: Institute of Applied Biotechnology, Biberach University of Applied Sciences, Hubertus-Liebrecht-Str. 35, 88400, Biberach, Germany.
authors.forename: Simon
authors.initials: S
authors.lastname: Fischer
authors.affiliation: Institute of Applied Biotechnology, Biberach University of Applied Sciences, Hubertus-Liebrecht-Str. 35, 88400, Biberach, Germany.
authors.forename: Fabian
authors.initials: F
authors.lastname: Stiefel
authors.affiliation: Genomics Core Facility, Medical Faculty, Ulm University, Ulm, Germany.
authors.forename: Karlheinz
authors.initials: K
authors.lastname: Holzmann
authors.affiliation: Institute of Applied Biotechnology, Biberach University of Applied Sciences, Hubertus-Liebrecht-Str. 35, 88400, Biberach, Germany.
authors.forename: René
authors.initials: R
authors.lastname: Handrick
authors.affiliation: Institute of Applied Biotechnology, Biberach University of Applied Sciences, Hubertus-Liebrecht-Str. 35, 88400, Biberach, Germany.
authors.forename: Friedemann
authors.initials: F
authors.lastname: Hesse
authors.affiliation: Rentschler Biotechnologie GmbH, Erwin-Rentschler-Str. 21, Laupheim, Germany.|VBBio Consultant, Auf dem Berg 17, Laupheim, Germany.
authors.forename: Markus
authors.initials: M
authors.lastname: Hörer
authors.affiliation: Department of Gene Therapy, Ulm University, Ulm, Germany.
authors.forename: Stefan
authors.initials: S
authors.lastname: Kochanek
authors.affiliation: Institute of Applied Biotechnology, Biberach University of Applied Sciences, Hubertus-Liebrecht-Str. 35, 88400, Biberach, Germany. otte@hochschule-bc.de.
authors.forename: Kerstin
authors.initials: K
authors.lastname: Otte
mesh_terms: D000818:Animals; D016466:CHO Cells; D002470:Cell Survival; D003412:Cricetulus; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation / Q000528:radiation effects*; D006367:HeLa Cells; D006801:Humans; D035683:MicroRNAs / Q000378:metabolism*; D011994:Recombinant Proteins / Q000096:biosynthesis*; D013696:Temperature
chemical_list: D035683:MicroRNAs; D011994:Recombinant Proteins
keywords: Adeno-associated virus; HeLa packaging cells; hypothermia; miR-483; microRNA; viral vector