Bistable Switches for Yeast Population Growth Designed on Predictions from a Toolkit of Cell Division Regulators

Positive feedback loop gene circuits are thought to control the process of cell differentiation. One such example is the cell-cycle lengthening mechanism that, through protein accumulation, leads to myeloid differentiation in stem cells [Kueh et al, 2013] and, the differentiation of neural precursors [Salomoni et al, 2010]. In both systems, a regulatory factor slows down cell division leading to its own accumulation, which constitutes de facto a positive feedback between the cell cycle and the factor itself. This naturally occurring principle has been exploited to construct a synthetic circuit that causes bistable gene expression in E. coli [Tan et al, 2009]. However, we propose to modulate the growth rate to design synthetic switches.

Here, we propose a library of parts to control/tune cell growth in yeast, and we show how we used it to rationally design a growth-based bistable switch. Through a literature search, we identified Yeast genes susceptible to induce growth arrest and use their antagonists to modulate the growth rate. Each gene was then expressed from an inducible promoter and resultant growth rates quantified. Finally, we used data-trained mathematical models to have a quantitative and qualitative description of each gene and their effects on cell division. We made use of this mathematical characterization to search for combinations of parts that leads to growth bistability. In a first attempt to build a growth-based bistable switch, we found that not only were the yeast cells were able to switch between the two states at any given time but also, the bistable switch remained in either state for over 5 days without loss of function.

In summary, we constructed and characterized a vocabulary of parts to control the growth rate of Yeast cells. We mathematically demonstrated that these parts could be used to systematically construct several growth-dependent behaviors in a predictable fashion. As a proof of concept, we designed and built the first stable growth-based bistable switch in yeast, and described a method that could be easily used to increase the number of these systems.