Triglyceride deficiency and diacylglycerol kinase1 activity lead to the upregulation of mevalonate pathway in yeast: A study for the development of potential yeast platform for improved production of triterpenoid

Besides energy storage and membrane biogenesis, lipids are known for their numerous biological functions. The two essential lipids, diacylglycerol (DG) and phosphatidic acid (PA), are shown to be associated with cell signalling processes. In this study, we examined whether triglyceride-deficient yeast mutants (tg Delta), dga1 Delta and dga1 Delta lro1 Delta, may play an important role in mevalonate (MEV) pathway regulation. Our metabolite analyses revealed that tg Delta cells showed high levels of squalene (SQ) and ergosterol (ERG), which are key indicators of MEV pathway activity. In addition, gene expression studies indicated that the MEV pathway genes in tg Delta cells were significantly upregulated. Interestingly, tg Delta cells exhibited high diacylglycerol kinase1 (DGK1) expression. Furthermore, DGK1 overexpression in WT and tg Delta phenotypes causes a substantial elevation in SQ and ERG levels, and we also found a significant increase in transcript levels of MEV pathway genes, confirming the new role of DGK1 in MEV pathway regulation. This suggests that high DG phosphorylation activity increases the PA pool that may induce the upregulation of MEV pathway in tg Delta cells. The induced MEV pathway is one of the key strategies in the field of synthetic biology for improved production of terpenoids in yeast. Thus, to examine whether increased endogenous MEV pathway flux can be redirected to triterpenoid, beta-Amyrin synthase gene was heterologously expressed in DGK1 overexpressing tg Delta cells that led to significant production of beta-Amyrin, a natural triterpenoid. In conclusion, our findings provide a novel strategy to increase MEV pathway precursors by modulating endogenous signal lipids for improved production of terpenoids.