A research group led by Graduate Student Akari Maeda and Professor Norihito Nakamichi of the Graduate School of Bioagricultural Sciences at Nagoya University has announced that they discovered a new molecular mechanism for the plant circadian clock, which deviates from the general rule of biochemical reactions in that it proceeds at a constant speed independent of ambient temperature. Degradation of the proteins TOC1 and PRR5, which have been known to play a role as a brake in the plant circadian clock, was found to occur at low temperatures, allowing the circadian clock to function in an ambient temperature-independent manner. The findings are expected to contribute to a better understanding of the temperature nociception systems of other species. The results were published in Science Advances on September 28.
Living organisms have an internal mechanism to measure an approximate circadian cycle (circadian clock) for accurately predicting diel changes in their environment. Biochemical reactions in the body proceed more rapidly as the temperature increases, but the circadian clock is practically unaffected by temperature. This property is called "temperature compensation of the circadian period length." The eukaryotic circadian clocks are thought to measure circadian rhythms by combining multiple biochemical reactions. However, the underlying molecular mechanism was unknown.
Using Arabidopsis thaliana, in which many circadian clock components have been found, the study aimed to identify the reactions required to drive the circadian clock, particularly those dependent on temperature. First, the research group introduced a reporter gene into the clock gene mutants to monitor the expression of the clock gene CCA1 and measured the period lengths of the circadian rhythm in the resulting mutants at temperatures ranging from 12℃ to 28℃. The result showed that the prr5 and toc1 mutants lost the temperature compensation most notably.
The speed of the clock increased with temperature in both mutants, suggesting that both genes act as a brake to slow the clock at high temperatures. To verify this function, they observed the strains overexpressing PRR5 and TOC1 and found that this trait became more pronounced at a higher temperature and disappeared at a low temperature. PRR5 and TOC1 proteins were shown to increase at high temperatures while they underwent ubiquitination and were actively degraded at low temperatures. Thus, in plants, temperature-dependent quantitative changes in PRR5 and TOC1 function as a brake that is activated at high temperatures.
Proteomics analysis to explore factors involved in regulating brake strength led to the discovery of LKP2, a ubiquitin E3 ligase that interacts with PRR5 and TOC1 in a cold-inducible manner. The lkp2 mutant exhibited extended periodicity at low temperatures with unnecessarily long circadian period lengths. The plant circadian clock is considered to be appropriately braked by LKP2, which regulates the amounts of PRR5 and TOC1 in response to ambient temperature so that the circadian clock pace remains constant irrespective of ambient temperature fluctuations.
Nakamichi said, "We were interested in the 'mechanism' by which the Arabidopsis clock resists temperature, but the reviewers of our paper asked, 'How can this mechanism be interpreted from an evolutionary perspective?' which we had never considered. We could not get a clear answer to this point of view in this study, but we are currently working on it as one of the next themes we want to solve."
Journal Information
Publication: Science Advances
Title: Cold-induced degradation of core clock proteins implements temperature compensation in the Arabidopsis circadian clock
DOI: 10.1126/sciadv.adq0187
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