Overexpression of the transcription factor Yap1 modifies intracellular redox conditions and enhances recombinant protein secretion
Authors:Marizela Delic1,2, Alexandra B. Graf2,3, Gunda Koellensperger1,4, Christina Haberhauer-Troyer1,4, Stephan Hann1,4, Diethard Mattanovich1,2, Brigitte Gasser1,2
doi: 10.15698/mic2014.11.173
Volume 1, pp. 376 to 386, published 31/10/2014.
1 Department of Biotechnology, BOKU University of Natural Resources and Life Sciences Vienna, Vienna, Austria.
2 Austrian Centre of Industrial Biotechnology (ACIB), Vienna, Austria.
3 School of Bioengineering, University of Applied Sciences FH Campus Wien, Vienna, Austria.
4 Department of Chemistry, BOKU University of Natural Resources and Life Sciences Vienna, Vienna, Austria.
Keywords:
ER, cytosol, cellular redox regulation, oxidative protein folding, glutathione, redox sensitive roGFP, Pichia pastoris.
Abbreviations:
ER - endoplasmic reticulum,
PDI - protein disulfide isomerase,
ROS - reactive oxygen species,
roGFP - redox sensitive variants of green fluorescent protein,
SOD - superoxide dismutase,
TRP - trypsinogen,
UPR - unfolded protein response.
Corresponding Author(s):
Conflict of interest statement:
The authors declare no conflict of interest.
Please cite this article as:
Marizela Delic, Alexandra B. Graf, Gunda Koellensperger, Christina Haberhauer-Troyer, Stephan Hann, Diethard Mattanovich, Brigitte Gasser (2014). Overexpression of the transcription factor Yap1 modifies intracellular redox conditions and enhances recombinant protein secretion. Microbial Cell 1(11): 376-386.
© 2014 Delic et al. This is an open-access article released under the terms of the Creative Commons Attribution (CC BY) license, which allows the unrestricted use, distribution, and reproduction in any medium, provided the original author and source are acknowledged.
Abstract:
Oxidative folding of secretory proteins in the endoplasmic reticulum (ER) is a redox active process, which also impacts the redox conditions in the cytosol. As the transcription factor Yap1 is involved in the transcriptional response to oxidative stress, we investigate its role upon the production of secretory proteins, using the yeast Pichia pastoris as model, and report a novel important role of Yap1 during oxidative protein folding. Yap1 is needed for the detoxification of reactive oxygen species (ROS) caused by increased oxidative protein folding. Constitutive co-overexpression of PpYAP1 leads to increased levels of secreted recombinant protein, while a lowered Yap1 function leads to accumulation of ROS and strong flocculation. Transcriptional analysis revealed that more than 150 genes were affected by overexpression of YAP1, in particular genes coding for antioxidant enzymes or involved in oxidation-reduction processes. By monitoring intracellular redox conditions within the cytosol and the ER using redox-sensitive roGFP1 variants, we could show that overexpression of YAP1 restores cellular redox conditions of protein-secreting P. pastoris by reoxidizing the cytosolic redox state to the levels of the wild type. These alterations are also reflected by increased levels of oxidized intracellular glutathione (GSSG) in the YAP1 co-overexpressing strain. Taken together, these data indicate a strong impact of intracellular redox balance on the secretion of (recombinant) proteins without affecting protein folding per se. Re-establishing suitable redox conditions by tuning the antioxidant capacity of the cell reduces metabolic load and cell stress caused by high oxidative protein folding load, thereby increasing the secretion capacity.