Multi-omics approaches to unravel regulatory dynamics in yeast bioreactor cultivations

Doctoral thesis, 2024

Climate change is a multifaceted problem that requires multiple scientific discoveries and engineering innovations. Among the innovations that have emerged in recent years are microbial cell factories, engineered microorganisms that produce desired molecules through their metabolism.

A promising microbial cell factory is Yarrowia lipolytica, an oleaginous yeast that has gained significant traction since it proved a versatile host to produce lipids as well as both bulk and fine chemicals. However, further research is needed to better understand this host and to design better bioprocesses.

To improve the current understanding of Y. lipolytica as a microbial cell factory, I combined chemostat cultivations with transcriptomic analysis. I studied the underlying biology of a platform strain with disrupted lipid synthesis, revealing that abolishing storage lipids induces protein misfolding and stress responses. I then explored the use of urea as an alternative and more sustainable nitrogen source, demonstrating that it does not alter the cell transcriptome and can reduce media acidification. I combined this information to improve a fed-batch cultivation to produce high titres of itaconic acid.

Meanwhile, I laid the foundations for single-cell transcriptomics to explore cell heterogeneity in bioreactor cultivations. I performed a proof-of-concept analysis in the well‑characterized yeast Saccharomyces cerevisiae to understand the potential challenges in translating single‑cell transcriptomics to Y. lipolytica. I found that cell cycle genes are a major source of variability that needs to be minimized.

The work performed combines bioreactor cultivation with omics analyses to inform and guide future strain improvement. Overall, this thesis explores and expands knowledge in relevant areas to develop Y. lipolytica as a microbial cell factory for the sustainable production of non‑lipid chemicals.