Analysis Of The Mechanism Underlying The Coordination Of Circadian Clock With Cold Response In Rice

Chilling stress is one of the main abiotic stresses that affect the growth,development and geographical distribution of rice.The research on the molecular mechanisms of rice response to chilling stress and regulation of cold tolerance has important theoretical significance for the breeding of cold resistant rice varieties.Circadian clock is an endogenous regulatory mechanism evolved by plants in the long-term adaptation to the alternation of day and night.The clock plays privatol roles in many important physiological processes and helps plants better adapt to the environment.The clock has a very promising application value in enhancing crop stress resistance,expanding the crop distribution and so on.However,it is not clear whether the chilling response of rice is regulated by circadian clock.In this study,we treated hybrid rice restorer 93-11 with chilling stress under the mimic natural light/dark cycles.It was found that the rice leaves suffered from chilling stress only at night showed the yellow and wilt leaves.Compared with the leaves suffered from the chilling stress only in the daytime,the chlorophyll content and photosynthetic efficiency were significantly reduced.Transcriptome analysis showed that the effects of low temperature stress on a considerable number of cold regulated genes(CORS)in different zeitberger time(ZT)were quite different.For example,KEGG functional analysis demonstrated that the genes in porphyrin and chlorophyll metabolism,photosynthesis antenna protein,photosynthesis and other physiological metabolic pathways were up-regulated by chilling stress in the daytime but down-regulated by chilling stress at night.Real-time quantitative PCR analysis further showed that chilling stress during several ZT time periods at night inhibited the expression of key enzymes encoding genes YGL8,Os PORA,LYL1 and Os CAO1,other genes for promoting chlorophyll accumulation such as SGRL and Hem A1,and the photosystem II antenna protein coding genes LHCB1 a and LHCB1 b,while chilling stress inmost ZT time periods during the day promoted their expression.The transcriptome analysis also found that a gene,OsSGL(stress and grain length),which has been reported to regulate rice yield and drought tolerance,was significantly induced by low temperature,and the induced amplitude was affected by ZT time period of chilling stress.In this study,it was found that the survival rate of OsSGL overexpressors in93-11 background was lower than that of wild type.It is suggested that the gene may be a major gene related to cold tolerance regulated by circadian clock.To further analyze the molecular regulatory mechanism of the gene on the biochemical level in the future,the physical and chemical properties of OsSGL protein were analyzed by bioinformatics,and His-tagged OsSGL protein was expressed and purified in prokaryotic expression system.For the first time,this study reveals the important role of circadian clock in regulating rice chilling responses by modulating chlorophyll level and photosynthesis efficiency.At least in part,the modulation may be through regulation the expression of chlorophyll metabolism and photosynthesis related genes.Further study of the target gene OsSGL,whose chilling sensitivity is regulated by circadian clock,negatively regulates chilling tolerance.Purification of OsSGL protein through the prokaryotic expression system will lay a foundation for further elucidation of the role of the gene as a potential molecular node for circadian clock to regulate the cold response in the future.