Collective behavior of oscillators in fungal filaments of Neurospora crassa

About This Grant

Collective behavior can be observed in a variety of contexts, the blinking of fireflies, the schooling of fish, the marching of locusts, and the flocking of birds. Collective behavior can also be observed at other levels of biological organization, such as the synchronized behavior of cells in tissues. Single cells, for example, have a biological clock, but their synchronized timekeeping is usually only observed at the level of tens of millions of cells. A grand challenge is understanding how do cellular clocks in cells and tissues become synchronized in the whole organism to keep time. The focus here will be on understanding the synchronization of cellular clocks in a model fungal system, Neurospora crassa. There are two principal theories for how clock synchronization arises; (1) through a shared biochemical signal between cellular clocks; (2) through a physical theory in which the random switching on and off of clock genes within a cell plays an essential role in synchronizing cellular clocks. In testing these two scientific theories we will come to understand the origin of the biological clock. Understanding the molecular origin of biological clocks could have broad implications, including engineering the timed delivery of therapeutics for improving human health, developing strategies to control agricultural pests like marching locusts, and timing bacterial assemblages in the world's oceans to impact carbon cycling in marine ecosystems. In order to understand the synchronization of clocks between cells, we face several challenges, including (1) lack of understanding on how the clock functions in the predominant life stage of the organism, the filament in its network; (2) lack of understanding how filaments in a network synchronize their clocks; (3) lack of models describing how the clock tells time in filaments. To answer these questions we will first develop microfluidics platforms (similar to integrated circuits for fluids) to make high-throughput and high-precision measurements on oscillators in living N. crassa filaments. Single filament measurements on single clock RNAs in this platform will enable us to confirm whether or not individual filaments have clocks and synchronize by a shared signal in the media called a quorum sensing signal or by direct contact. We will better understand the microfluidic experiments with a physical model of the clock in growing filaments by measurements derived from the microfluidic devices. We have developed a novel NMR methodology called Continuous in vivo Metabolism-NMR or CIVM-NMR allowing real time measurement of metabolites in living cells to link metabolites in the cell with their binding proteins involved in cellular clock synchronization in the same spirit that the Nobel Laureates, Beadle and Tatum, were able to link genes with proteins in metabolism. An interdisciplinary team from genetics, engineering, physics and chemistry will tackle this challenge. We will develop novel microfluidics platforms to measure phase synchronization of biological clocks in living N. crassa filaments. We will develop and test novel clock models in filaments using novel ensemble methods from Statistical Physics. We will also identify and test signaling molecules and their associated proteins that may be responsible for filament synchronization by a newly developed method of CIVM-NMR and fractionating metabolites into libraries containing different synchronization signals for testing from Chemistry. We expect to achieve our objective by pursuing three tasks: (1) confirming whether or not individual filaments have clocks and synchronize by quorum sensing or by contact; (2) understanding the microfluidic experiments via a physical model of the clock in filaments specified by the measurements in task 1; (3) linking metabolites and associated proteins in the cell with clock signaling for cellular clock synchronization. Education and outreach activities open to everyone are designed around the research focus of this project, biological clocks and their synchronization. (1) An interdisciplinary research-centered course, Clock Collaboratorium, will be developed. (2) Undergraduate research projects will be developed with the central theme of the biological clock, and used in two NSF REU site programs. (3) We will help build a collective behavior community through a new Research Conference as done previously in a Gordon Research Conference. This project is supported by the Systems and Synthetic Biology cluster within the Division of Molecular and Cellular Biosciences. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.