Chemical Perturbation of Chloroplast-Related Processes Affects Circadian Rhythms of Gene Expression in Arabidopsis: Salicylic Acid Application Can Entrain the Clock.

The plant circadian system reciprocally interacts with metabolic processes. To investigate entrainment features in metabolic-circadian interactions, we used a chemical approach to perturb metabolism and monitored the pace of nuclear-driven circadian oscillations. We found that chemicals that alter chloroplast-related functions modified the circadian rhythms. Both vitamin C and paraquat altered the circadian period in a light-quality-dependent manner, whereas rifampicin lengthened the circadian period under darkness. Salicylic acid (SA) increased oscillatory robustness and shortened the period. The latter was attenuated by sucrose addition and was also gated, taking place during the first 3 h of the subjective day. Furthermore, the effect of SA on period length was dependent on light quality and genotype. Period lengthening or shortening by these chemicals was correlated to their inferred impact on photosynthetic electron transport activity and the redox state of plastoquinone (PQ). Based on these data and on previous publications on circadian effects that alter the redox state of PQ, we propose that the photosynthetic electron transport and the redox state of PQ participate in circadian periodicity. Moreover, coupling between chloroplast-derived signals and nuclear oscillations, as observed in our chemical and genetic assays, produces traits that are predicted by previous models. SA signaling or a related process forms a rhythmic input loop to drive robust nuclear oscillations in the context predicted by the zeitnehmer model, which was previously developed for Neurospora. We further discuss the possibility that electron transport chains (ETCs) are part of this mechanism.

Physiological and Genetic Dissection of Sucrose Inputs to the Arabidopsis thaliana Circadian System.

Circadian rhythms allow an organism to synchronize internal physiological responses to the external environment. Perception of external signals such as light and temperature are critical in the entrainment of the oscillator. However, sugar can also act as an entraining signal. In this work, we have confirmed that sucrose accelerates the circadian period, but this observed effect is dependent on the reporter gene used. This observed response was dependent on sucrose being available during free-running conditions. If sucrose was applied during entrainment, the circadian period was only temporally accelerated, if any effect was observed at all. We also found that sucrose acts to stabilize the robustness of the circadian period under red light or blue light, in addition to its previously described role in stabilizing the robustness of rhythms in the dark. Finally, we also found that CCA1 is required for both a short- and long-term response of the circadian oscillator to sucrose, while LHY acts to attenuate the effects of sucrose on circadian period. Together, this work highlights new pathways for how sucrose could be signaling to the oscillator and reveals further functional separation of CCA1 and LHY.

HSP90 Contributes to Entrainment of the Arabidopsis Circadian Clock via the Morning Loop.

The plant circadian clock allows the synchronization of internal physiological responses to match the predicted environment. HSP90.2 is a molecular chaperone that has been previously described as required for the proper functioning of the Arabidopsis oscillator under both ambient and warm temperatures. Here, we have characterized the circadian phenotype of the hsp90.2-3 mutant. As previously reported using pharmacological or RNA interference inhibitors of HSP90 function, we found that hsp90.2-3 lengthens the circadian period and that the observed period lengthening was more exaggerated in warm-cold-entrained seedlings. However, we observed no role for the previously identified interactors of HSP90.2, GIGANTEA and ZEITLUPPE, in HSP90-mediated period lengthening. We constructed phase-response curves (PRCs) in response to warmth pulses to identify the entry point of HSP90.2 to the oscillator. These PRCs revealed that hsp90.2-3 has a circadian defect within the morning. Analysis of the cca1, lhy, prr9, and prr7 mutants revealed a role for CCA1, LHY, and PRR7, but not PRR9, in HSP90.2 action to the circadian oscillator. Overall, we define a potential pathway for how HSP90.2 can entrain the Arabidopsis circadian oscillator.