Positive genetic interactors of HMG2 identify a new set of genetic perturbations for improving sesquiterpene production in Saccharomyces cerevisiae.

Terpenoids and isoprenoids are an important class of natural products, which includes currently used drugs, high value bioactive and industrial compounds, and fuel candidates. Due to their industrial application, there is increasing interest in the development of S. cerevisiae strains capable of producing high levels of terpenoids. Aiming to identify new gene targets which can be manipulated to increase sesquiterpene production, a set of HMG2 positive genetic interactors were assessed as single and digenic heterozygous deletions in the presence or absence of stable HMG2(K6R) overexpression. Upon single allele deletion, most genes examined led to increased sesquiterpene production in yeast cells. Tandem heterozygous deletion of a set of three genes, the ubiquitin ligases ubc7 and ssm4/doa10, and the ER resident protein pho86, led to an 11-fold increase in caryophyllene yields (125 mg/L in shake flasks) compared to cells lacking these modifications. The effect of the heterozygous deletions appears to be due to Hmg1p and Hmg2p stabilization. Heterozygous deletions cause significant reductions in protein levels but do not lead to growth impediments frequently seen in haploid strains. By exploiting desirable haploinsufficiencies in yeast, we identified a new set of genes that can be disrupted in tandem and cause significant stabilization of Hmgp and a substantial increase in sesquiterpene production. The approach presented here allows new genetic perturbations to be compiled on yeast cell factory strains without negatively impacting cell growth and viability.