Harnessing the yeast nucleus for the efficient production of valuable monoterpenes

Subcellular engineering emerges as a pivotal strategy to amplify terpene production in yeast cell factories. The yeast nucleus, which shares acetyl-CoA pool with the cytosol and maintains nuclear membrane integrity during mitosis, offers unique compartmentalization capabilities. Despite these attributes, there is a notable absence of research harnessing the potential of the yeast nucleus for subcellular engineering. In this study, we developed the yeast nucleus as a novel platform, capitalizing on the benefits of subcellular compartments to efficiently produce valuable chemicals. Microbial production of monoterpenes remains a formidable obstacle due to the inherent coupling of the precursor geranyl diphosphate with cytosolic isoprenoid biosynthesis. The introduction of the mevalonate pathway for effective utilization of the acetyl-CoA pool in the nucleus resulted in comparable production levels of several commercially interesting monoterpenes. Furthermore, we demonstrate that combining nuclear engineering with other subcellular compartments synergistically increases production. Through combinatorial modification of the nucleus and peroxisomes, the titer of (R)-(+)-limonene production was increased by 303% compared to only using peroxisomes alone, reaching 362.07 mg/L during flask-batch fermentation from 2% galactose. This study demonstrates that the nucleus can be a powerful platform for the efficient production of valuable compounds, particularly those derived from acetyl-CoA.