Project outline

Background

The circadian clock is a gene circuit that generates 24-h cycles of gene expression in many organisms in anticipation of daily cycles of sunlight and temperature. In the cyanobacterium S. elongatus, which has the simplest clock we know of, most of the genome is under clock control. The cyanobacterial clock regulates many processes inside the cell, including metabolism, photosynthesis or the cell cycle. While the clock anticipates environmental change, it is also influenced by a variety of signalling cues. Light and temperature transitions reset the clock state, but the effect of these signals on clock proteins is relayed through more fundamental quantities, such as redox metabolites and ATP/ADP ratios. These cues can be said to characterise the global energy state of the cell, but on the other hand the clock controls expression of most photosynthetic and metabolic proteins. This dynamic and bi-directional coupling between the circadian clock and metabolism likely holds the key to understanding the evolutionary significance of clocks at the most fundamental level (1, 2).

General aim and methodology

The aim of this project is to characterise the cross-talk between the circadian clock and metabolism and understand how this cross-talk coordinates cellular growth and division under different environmental scenarios. Using fluorescent reporters, the joint dynamics of the clock and metabolic enzymes will be measured using time-lapse microscopy (3) and microfluidics for thousands of individual cells. Cellular energetics (e.g. ATP/ADP ratios) will be monitored through ratiometric fluorescent reporters. These measurements will provide a comprehensive timeline of physiological changes throughout the 24-h cycle and of how cells allocate metabolic resources to time growth, cell division and DNA replication, which will allow us to create an integrated model of the coupling between the clock and metabolism. We will test this model using a combination of genetic and environmental perturbations in microfluidic devices. These results will allow us to understand why clocks offer a selective advantage, and thus reveal a mechanistic ‘logic’ that can be applied to other systems.

References

A. M. Puszynska, E. K. O’Shea, Switching of metabolic programs in response to light availability is an essential function of the cyanobacterial circadian output pathway. eLife 6, e23210 (2017).

G. Pattanayak, M. J. Rust, The Cyanobacterial Clock and Metabolism. Curr Opin Microbiol 18, 90–95 (2014).

B. M. Martins, A. K. Das, L. Antunes, J. C. Locke, Frequency doubling in the cyanobacterial circadian clock. Molecular Systems Biology12, 896 (2016).

Techniques

• Single-cell time-lapse microscopy and microfluidics
• Computational work including quantitative image analysis, time-series analysis, writing short routines for automated microscopy acquisition and possibly modelling (depending on the background and interests of the student)
• Microbiology and molecular genetics