Abstract
In this research, we were interested in answering a question whether subjecting a Yarrowia lipolytica strain overproducing a recombinant secretory protein (rs-Prot) to pre-optimized stress factors may enhance synthesis of the rs-Prot. Increased osmolarity (3 Osm kg−1) was the primary stress factor implemented alone or in combination with decreased temperature (20 °C), known to promote synthesis of rs-Prots. The treatments were executed in batch bioreactor cultures, and the cellular response was studied in terms of culture progression, gene expression and global proteomics, to get insight into molecular bases underlying an awaken reaction. Primarily, we observed that hyperosmolarity executed by high sorbitol concentration does not enhance synthesis of the rs-Prot but increases its transcription. Expectedly, hyperosmolarity induced synthesis of polyols at the expense of citric acid synthesis and growth, which was severely limited. A number of stress-related proteins were upregulated, including heat-shock proteins (HSPs) and aldo–keto reductases, as observed at transcriptomics and proteomics levels. Concerted downregulation of central carbon metabolism, including glycolysis, tricarboxylic acid cycle and fatty acid synthesis, highlighted redirection of carbon fluxes. Elevated abundance of HSPs and osmolytes did not outbalance the severe limitation of protein synthesis, marked by orchestrated downregulation of translation (elongation factors, several aa-tRNA synthetases), amino acid biosynthesis and ribosome biogenesis in response to the hyperosmolarity. Altogether we settled that increased osmolarity is not beneficial for rs-Prots synthesis in Y. lipolytica, even though some elements of the response could assist this process. Insight into global changes in the yeast proteome under the treatments is provided.Key points• Temp enhances, but Osm decreases rs-Prots synthesis by Y. lipolytica.• Enhanced abundance of HSPs and osmolytes is overweighted by limited translation.• Global proteome under Osm, Temp and Osm Temp treatments was studied.
Highlights
Environmental factors evoke multilayered responses in the yeast producer cells
Maturation and secretion of recombinant secretory protein (rs-Prot) is a complex biological process relying on multiple molecular functions
High provision of heat-shock proteins (HSPs) and osmolytes within the producer cell seems to be a potentially generalizable strategy for enhancing rs-Prot production in bacteria and yeast (Oganesyan et al 2007; Lazar et al 2011). It is a widely recognized mechanism, that under exposure to different environmental stress factors, the cell increases the intracellular abundance of the protective agents to preserve delicate proteins and membranes from denaturation
Summary
Environmental factors evoke multilayered responses in the yeast producer cells. Some treatment conditions are known to promote synthesis of specific compounds, while the others exert adverse impact in this regard. It is widely recognized that hyperosmotic conditions promote synthesis of polyols in yeast and that decreased temperature is beneficial for synthesis and secretion of recombinant, secretory proteins (rs-Prots). Both these phenomena were observed and employed in bioprocesses with a nonconventional yeast species Yarrowia lipolytica (Tomaszewska et al 2012; Yang et al 2014, 2015; Kubiak et al 2021),. We are interested in answering a question whether subjecting a Y. lipolytica production culture to a specific, pre-optimized stress factor may enhance synthesis of rs-Prots. While the relationship between osmotic stress and enhanced synthesis of polyols in Y. lipolytica is clearly understood and well-described at molecular level (Kobayashi et al 2015; Rzechonek et al 2018), the effect of the former on the production of rs-Prots requires further studies
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