Abstract

Production of biopharmaceuticals relies on the expression of mammalian cDNAs in host organisms. Here we show that the expression of a human cDNA in the moss Physcomitrium patens generates the expected full-length and four additional transcripts due to unexpected splicing. This mRNA splicing results in non-functional protein isoforms, cellular misallocation of the proteins and low product yields. We integrated these results together with the results of our analysis of all 32,926 protein-encoding Physcomitrella genes and their 87,533 annotated transcripts in a web application, physCO, for automatized optimization. A thus optimized cDNA results in about twelve times more protein, which correctly localizes to the ER. An analysis of codon preferences of different production hosts suggests that similar effects occur also in non-plant hosts. We anticipate that the use of our methodology will prevent so far undetected mRNA heterosplicing resulting in maximized functional protein amounts for basic biology and biotechnology.

Top et al. report the expression of a human blood-clotting factor IX-encoding cDNA in the moss Physcomitrium patens and demonstrate that in addition to the unspliced full-length transcript, the splicing machinery produced several different transcripts, and thus different protein isoforms, which the authors designate as heterosplicing. The authors present a web application, physCO, to automatize the process of P. patens-specific codon optimization, a tool that can assist in maximized functional protein amounts for biotechnology.

Details

Title
Expression of a human cDNA in moss results in spliced mRNAs and fragmentary protein isoforms
Author
Top Oguz 1   VIAFID ORCID Logo  ; Milferstaedt Stella W L 2   VIAFID ORCID Logo  ; van Gessel Nico 3   VIAFID ORCID Logo  ; Hoernstein Sebastian N W 3   VIAFID ORCID Logo  ; Özdemir Bugra 3   VIAFID ORCID Logo  ; Decker, Eva L 3   VIAFID ORCID Logo  ; Reski Ralf 4   VIAFID ORCID Logo 

 University of Freiburg, Plant Biotechnology, Faculty of Biology, Freiburg, Germany (GRID:grid.5963.9); University of Freiburg, Spemann Graduate School of Biology and Medicine (SGBM), Freiburg, Germany (GRID:grid.5963.9); Ludwig-Maximilians-University Munich, Plant Molecular Cell Biology, Department Biology I, LMU Biocenter, Planegg-Martinsried, Germany (GRID:grid.5252.0) (ISNI:0000 0004 1936 973X) 
 University of Freiburg, Plant Biotechnology, Faculty of Biology, Freiburg, Germany (GRID:grid.5963.9); University of Freiburg, Cluster of Excellence livMatS @ FIT – Freiburg Center for Interactive Materials and Bioinspired Technologies, Freiburg, Germany (GRID:grid.5963.9) 
 University of Freiburg, Plant Biotechnology, Faculty of Biology, Freiburg, Germany (GRID:grid.5963.9) 
 University of Freiburg, Plant Biotechnology, Faculty of Biology, Freiburg, Germany (GRID:grid.5963.9); University of Freiburg, Spemann Graduate School of Biology and Medicine (SGBM), Freiburg, Germany (GRID:grid.5963.9); University of Freiburg, Cluster of Excellence livMatS @ FIT – Freiburg Center for Interactive Materials and Bioinspired Technologies, Freiburg, Germany (GRID:grid.5963.9); CIBSS – Centre for Integrative Biological Signalling Studies, Freiburg, Germany (GRID:grid.5963.9) 
Publication year
2021
Publication date
2021
Publisher
Nature Publishing Group
e-ISSN
23993642
Source type
Scholarly Journal
Language of publication
English
ProQuest document ID
2560479862
Copyright
© The Author(s) 2021. This work is published under http://creativecommons.org/licenses/by/4.0/ (the “License”). Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License.