Using systems biology to determine how budding yeast coordinates carbon and nitrogen sensing for efficient growth

A hallmark of life is the ability to sense and respond to change, and even unicellular organisms have this ability. They are able to sense changes in the availability of nutrients and adjust the rate at which they grow to new environments. Although we know the proteins involved in making this choice, we do not understand how they interact to enable cells to "decide" appropriately. A difficulty is monitoring the proteins as they perform decision-making. We will use bakers' yeast, a cell with a nucleus like our own cells, where we can measure its decision-making activity by the shuttling of proteins in and out of the nucleus. By exposing cells to changing levels of carbon and nitrogen sources - two essential nutrients, and by transiently removing proteins that are key to signalling, we will characterise how sensing is coordinated with growth. Through integrating this data with mathematical modelling, we will infer the decision-making strategy that is most supported by our results. Evolution means that yeast cells are related to human cells, and signalling proteins in yeast are similar to our own signalling proteins. This task of coordinating growth with the availability nutrients is fundamental, and the strategy we uncover should therefore be general and operate too in other organisms.

A key ability of cells is to coordinate sensing of extracellular carbon and nitrogen to enable efficient growth. Although we know that this ability comes from conserved intracellular molecules, how these molecules interact to generate decision-making is unclear. Research is hindered by the difficulty of measuring the activities of signalling molecules over time as cells respond. Using budding yeast, we will combine time-lapse microscopy with mathematical modelling to uncover the strategy they use for carbon and nitrogen sensing. To overcome the lack of real-time reporters, we will exploit that for yeast changes in extracellular nutrients cause tens of transcription factors to move into or out of the nucleus, with each transcription factor therefore a potential reporter once tagged with a fluorescent protein. With novel microfluidics, we will identify new translocating reporters for carbon and nitrogen sensing through a systematic single-cell study of almost all yeast's transcription factors. We will further develop a novel scalable technique that uses these reporters and transient perturbations to measure kinase and phosphatase activities over time in changing environments. Growth rate correlates with fitness in yeast, and we will use growth rate as a global output of signalling. Concurrently measuring growth rate and the activities of key molecules such as TOR and AMP kinase via the transcription factors, we will systematically characterise signalling in dynamic levels of extracellular carbon and nitrogen. To uncover how the molecules interact to enable growth, we will use mathematics to infer from our data the strategy cells use. This task, of deciding how fast to grow in changing availabilities of carbon and nitrogen, is fundamental and ancient, and the principles and the logic of signalling we discover should therefore impact widely.