Single Cells of Neurospora Crassa Show Circadian Oscillations, Light Entrainment, Temperature Compensation, and Phase Synchronization

Zhaojie Deng,Xiao Qiu,Brooke Hull,Michael T Judge,Leidong Mao,Heinz-Bernd Schuttler,Jia Hwei Cheong,Jonathan Arnold,James Griffith,Ahmad Al-Omari,Lingyun Wu,Cristian Caranica,Carmen Rodriguez,Sam Arsenault

doi:10.1109/access.2019.2910731

Abstract

Using a microfluidics device, fluorescence of a recorder (mCherry or mVenus) gene driven by a clock-controlled gene-2 promoter (ccg-2p) was measured simultaneously on over 1000 single cells of Neurospora crassa every half hour for 10 days under each of varied light and temperature conditions. Single cells were able to entrain to light over a wide range of day lengths, including 6, 12, or 36 h days. In addition, the period of oscillations in fluorescence remained remarkably stable over a physiological range of temperatures from 20 °C to 30 °C (Q 10 = 1.00-1.07). These results provide evidence of an autonomous clock in most single cells of N. crassa. While most cells had clocks, there was substantial variation between clocks as measured by their phase, raising the question of how such cellular clocks in single cells phase-synchronize to achieve circadian behavior in eukaryotic systems at the macroscopic level of 10 7 cells, where most measurements on the clock are performed. Single cells were placed out of phase by allowing one population to receive 6 or 12 h more light before lights out (D/D). The average phase difference was reduced in the mixed population relative to two unmixed control populations.

Highlights

The synchronization of the collective behavior of organisms can be observed in a variety of contexts [1], the blinking of fireflies, the collective marching of an army of locusts [2], the foraging of baboon troops [3], the schooling of fish, and the flocking of birds [4]
Light entrainment studies at the single cell level have included a small number of cells in Zebrafish [38], tracking of cells in the suprachiasmatic nuclei (SCN) of mice [19], tracking ∼100 single cells in tissue of the duckweed, Lemna gibba [39], and tracking ∼100 single cells in tissue of Arabidopsis thaliana [40]
Did we wish to test whether light entrainment occurs in single cells, we wanted to know whether single cells would track a L/D cycle over very short or very long days [19]

Summary

Introduction

The synchronization of the collective behavior of organisms can be observed in a variety of contexts [1], the blinking of fireflies, the collective marching of an army of locusts [2], the foraging of baboon troops [3], the schooling of fish, and the flocking of birds [4]. How this synchronization occurs remains a fundamental problem in biology. These biological clocks are observed at the level of 107 cells or more [6], where there is tight synchronization of the circadian rhythm between cells

Methods

Results

Discussion

Conclusion