Engineering biosensors for the detection of unfolded protein response in saccharomyces cerevisiae
thesisposted on 2022-03-28, 02:36 authored by Kai Peng
The unfolded protein response (UPR) is activated when the protein folding capacity of the endoplasmic reticulum (ER) is overwhelmed by the accumulation of unfolded proteins inside the ER lumen. In S. cerevisiae, the UPR facilitates the up-and down-regulations of a collection of UPR target genes via the IRE1/HAC1 dependent signalling pathway to bring the ER back to homeostasis. In the application of S. cerevisiae as a cell factory for producing heterologous proteins,low levels of target protein secretion are often associated with the up-regulation of UPR. In this study we constructed nine UPR biosensors using the putative promoter sequences of nine UPR genes including DER1, ERO1, EUG1, HAC1, KAR2, PDI1, PMT2, SEC12 andOST2as sensors, and green fluorescent protein as reporter. We have evaluated these biosensors in their capacities to detect the activation and different levels of UPR, and found that the sensors incorporating the DER1and ERO1 putative promoter sequences gave the best responses across a range of UPR induction levels. This study highlighted the prospect of providing a real-time and high-throughput detection of unfolded protein induced stress and its concomitant influences on protein secretion.