Abstract
Cellulosic plant biomass is a promising sustainable resource for generating alternative biofuels and biochemicals with microbial factories. But a remaining bottleneck is engineering microbes that are tolerant of toxins generated during biomass processing, because mechanisms of toxin defense are only beginning to emerge. Here, we exploited natural diversity in 165 Saccharomyces cerevisiae strains isolated from diverse geographical and ecological niches, to identify mechanisms of hydrolysate-toxin tolerance. We performed genome-wide association (GWA) analysis to identify genetic variants underlying toxin tolerance, and gene knockouts and allele-swap experiments to validate the involvement of implicated genes. In the process of this work, we uncovered a surprising difference in genetic architecture depending on strain background: in all but one case, knockout of implicated genes had a significant effect on toxin tolerance in one strain, but no significant effect in another strain. In fact, whether or not the gene was involved in tolerance in each strain background had a bigger contribution to strain-specific variation than allelic differences. Our results suggest a major difference in the underlying network of causal genes in different strains, suggesting that mechanisms of hydrolysate tolerance are very dependent on the genetic background. These results could have significant implications for interpreting GWA results and raise important considerations for engineering strategies for industrial strain improvement.
Highlights
The increased interest in renewable energy has focused attention on non-food plant biomass for the production of biofuels and biochemicals [1]
We identified 486,302 high quality Single nucleotide polymorphisms (SNPs). 68% of them had a minor allele frequency less than 5%
Most of these were from natural environments (11 strains) but they included clinical samples (5 strains), baking strains (3 strains), a sugar cane fermenter (1) and a laboratory strain (FL100, which was scored as 98% heterozygous and may have mated with another strain in its recent history (S1 Table))
Summary
The increased interest in renewable energy has focused attention on non-food plant biomass for the production of biofuels and biochemicals [1]. The concentrations and composition of these inhibitors vary for different pretreatment methods and depend on the plant feedstocks [7, 9, 11] These toxins decrease cell productivity by generating reactive oxygen species, damaging DNA, proteins, cell membranes [12,13,14], and inhibiting important physiological processes, including enzymes required for fermentation [15], de novo nucleotide biosynthesis [16], and translation [17]. Despite knowledge of these targets, much remains to be learned about how the complete suite of hydrolysate toxins (HTs) acts synergistically to inhibit cells. How the effects of HTs are compounded by other industrial stresses such as high osmolarity, thermal stress, and end-product toxicity remains murky
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