This paper reports the use of luciferase reporter gene imaging to monitor circadian rhythms in leaves of Arabidopsis thaliana plants, achieving resolution close to the cellular level. Leaves grown without environmental cycles for up to 3 wk reproducibly showed spatiotemporal waves of gene expression consistent with intercellular coupling, using several reporter genes. Within individual leaves, different regions differed in phase by up to 17 h. A broad range of patterns was observed among leaves, rather than a common spatial distribution of circadian properties. Leaves exposed to light–dark cycles always had fully synchronized rhythms, which could desynchronize rapidly. After 4 d in constant light, some leaves were as desynchronized as leaves grown without any rhythmic input. Applying light–dark cycles to such a leaf resulted in full synchronization within 2–4 d. Thus, the rhythms of all cells were coupled to external light–dark cycles far more strongly than the cellular clocks were coupled to each other. Spontaneous desynchronization under constant conditions was limited, consistent with weak intercellular coupling among heterogeneous clocks. Both the weakness of coupling and the heterogeneity among cells are relevant to interpret molecular studies and to understand the physiological functions of the plant circadian clock.
Benedicte Wenden, David L. K. Toner, Sarah K. Hodge, Ramon Grima, Andrew J. Millar (2012) Proc. Natl. Acad. Sci. USA, 109: 6757-62