| The rotation of the earth produces the alternation of the sun and the moon,and the long history of life has marked the rhythms of almost all organisms.The endogenous clock system,which emerged during the evolution of biological adaptations,penetrates into every aspect of the organism along with the beginning of life,and the periodic transcriptional regulation mechanism ensures that the organism can coordinate the external environmental changes in real time.A series of physiological and biochemical processes in plant growth,such as germination,flowering and fruiting,are inseparable from the material basis of support,but there is still a lack of comprehensive recognition of the effect of the rhythm system on metabolic levels.As a major food crop,the growth and developmental processes of rice have a close influence on the seeds and yield.The analysis of rice rhythmic metabolism will help us to deepen our understanding of the adaptive mechanisms during rice growth.In this study,metabolome and transcriptome material of leaf tissues was collected for two consecutive days for rhythmical studies by using four different indica-japonica varieties of rice accessions(MH63,ZS97,ZH11 and DJ).By combining targeted and untargeted metabolic detection methods,900 metabolites were quantitatively measured,and a large group of rhythmic metabolites were discovered,and more than a quarter of them showed the same rhythmicity among all four rice varieties,which showed great rhythmic differences between day and night.Amino acids,nucleotides,organic acids and carbohydrates were significantly enriched mainly in the afternoon,and vitamins and hormones fluctuated to their highest levels late at night,while lipids tended to reach their maximum accumulation at dawn.And based on the rhythm analysis of the transcriptome,more than 40% of the genes were found to be rhythmical in all four rice accessions.Compared with the metabolome,the fluctuating phase of the transcriptome transitioned smoothly from morning to the following dawn,fully demonstrating the fine regulation of transcriptional activities by the rhythm clock,with the main rhythmic transcriptional climax occurring in the period from evening to nightfall.The enrichment analysis of the rhythm genes showed that the main regulated metabolic pathways included amino acid,nucleotide and nicotinic acid pathways,which highly correspond to the enrichment results of metabolites.The rhythmic regulation pattern of rice metabolism was investigated by associating metabolic phenotypes with transcriptome results.We found that the transmission of rhythms in the carbohydrate and nucleotide metabolic pathways in rice is highly real-time,with changes in rhythm fluctuations occurring almost simultaneously.And subsequently,the temporal association analysis of rhythm dynamics revealed that the transcriptional and metabolic correlations were highest when the two had a 4-hour time difference,which was significantly higher than other time delays.By analyzing the time delay relationship between transcription and metabolism of JA and ABA in different manners,we confirmed the main4-hour delay time of rhythm from transcription transmitted to metabolism.Moreover,in the metabolic pathway,the difference between the phase of expression peaks of upstream and downstream related genes and the accumulation peak phase of metabolic terminal products gradually shortened from top to bottom,fully demonstrating the fine and flexible regulation of metabolic rhythms in transcriptional regulation.In addition,highly conserved metabolic rhythms maintain a coordinated metabolic homeostasis,but beyond the conserved metabolic rhythms,some metabolites such as branched-chain amino acids(BCAAs)display differences in rhythms among different rice varieties.We analyzed the mechanism of the formation of rhythmic differences in branchedchain amino acids as an example,and found a gene branched-chain amino acid transaminase 2(Os BCAT2)in its metabolic pathway.This enzyme can be involved in the transamination of branched-chain amino acids during the synthesis and deamination during degradation,and can have a direct effect on the changes in the content of BCAAs.The expression of Os BCAT2 was nearly 4-fold higher at dawn in japonica than in indica,and when it was mutated in japonica accession ZH11,the rhythmic fluctuation in the content of BCAAs was lost.It is suggested that it is the differential expression of Os BCAT2 at dawn that causes the diversity of BCAAs accumulation.In summary,by integrating the rice metabolome and transcriptome,we established a rhythm-based transcription-metabolism network and revealed the rhythm transmission pattern between the rice transcriptome and metabolome.Furthermore,our study demonstrated the conserved and diverse metabolic rhythms in different rice accessions,and focused on resolving the molecular mechanism of rhythmic accumulation diversity of branched-chain amino acids,which provides a new idea to utilize the rhythmic diversity among rice accessions to optimize growth and development regulation and improve nutritional quality. |
