Emergence of Phenotypically Distinct Subpopulations Is a Factor in Adaptation of Recombinant Saccharomyces cerevisiae under Glucose-Limited Conditions.

Cells cultured in a nutrient-limited environment can undergo adaptation, which confers improved fitness under long-term energy limitation. We have shown previously how a recombinant Saccharomyces cerevisiae strain, producing a heterologous insulin product, under glucose-limited conditions adapts over time at the average population level. Here, we investigated this adaptation at the single-cell level by application of fluorescence-activated cell sorting (FACS) and showed that the following three apparent phenotypes underlie the adaptive response observed at the bulk level: (i) cells that drastically reduced insulin production (23%), (ii) cells with reduced enzymatic capacity in central carbon metabolism (46%), and (iii) cells that exhibited pseudohyphal growth (31%). We speculate that the phenotypic heterogeneity is a result of different mechanisms to increase fitness. Cells with reduced insulin productivity have increased fitness by reducing the burden of the heterologous insulin production, and the populations with reduced enzymatic capacity of the central carbon metabolism and pseudohyphal growth have increased fitness toward the glucose-limited conditions. The results highlight the importance of considering population heterogeneity when studying adaptation and evolution.