Regulatory Mechanism Study Of The Linkage Between ?-aminobutyric Acid Metabolism And Leaf Rhythmic Movement In Arabidopsis Thaliana

The innate circadian clock in plant regulates the rhythmic expression of genes to adapt the plant growth and development to the outside constant varying environment.The biological rhythm output pathway at molecular level is characterized by rhythmic changes of gene expression and metabolites,and the external physiological processes such as leaf movement,stomatal conductance,flowering time determination and so on.The rhythmic leaf movement is a reliable external index for monitoring the internal rhythm regulation of the biological clock.More recent studies have shown that there is a mutual regulatory interplay between the rhythms of the biological clock and the metabolites.?-aminobutyric Acid?GABA?is an important metabolite in plants,which is widely involved in plant biotic and abiotic stresses and plant pollen fertilization.To elucidate the relationship between GABA and circadian clock regulation,using rhythmic leaf movement of Arabidopsis as monitoring indicators,the mutants of GABA biosynthesis and catabolism enzymes?glutamic acid decarboxylase,GAD;GABA transaminase,GABA-T/POP2;glutamate dehydrogenase,GDH,etc.?and the mutants of core clock gene?cca1,lhy and toc1,etc.?were employed to discuss the inherent interaction of GABA metabolism and biological rhythm.The results herein are as follows:1.Exogenous GABA(1.0 mmol·L^-1)could increase the amplitude of the rhythmic movement of the leaves in three ecotypes of Arabidopsis thaliana?Col,Ler and Ws?to different extent,and the amplitude had obvious changes in Col ecotype and Ws ecotype;2.Glutamate decarboxylase?GAD?is the key enzyme to control the biosynthesis of GABA.In GABA biosynthetic mutant?gad1-3?and double mutant of gad1/gad2,the amplitude of rhythmic movement of leaves is lower than that of wild type in different degrees,and the amplitude of leaf movement varies significantly in gad2 and gad1/gad2;3.GABA transaminase?GABA-T/POP2?is a key enzyme to catalyze GABA catabolism.Glutamate dehydrogenase?GDH?plays an important role in catalyzing the conversion from?-ketoglutaric acid,the intermediate metabolite of tricarboxylic acid cycle?TCA?,to glutamate acid,the precursor of GABA biosynthesis.In pop2 mutant,the amplitude of rhythmic change of leaf movement was significantly lower than that of control.In gdh1/gdh2 mutant,leaf movement showed arrhythmic pattern;4.CCA1,LHY and TOC1 are the central core elements to maintain the circadian rhythms of Arabidopsis thaliana.In the germination growth stage of Arabidopsis,the addition of exogenous GABA could weaken the level of LHY expression,but enhance the level of TOC1 expression.In the flowering stage of Arabidopsis,the addition of exogenous GABA has little effect on the level of CCA1,LHY and TOC1 expression.In cca1,lhy and toc1 mutants,leaf movement showed arrhythmic change pattern,but exogenous GABA addition could increase the amplitude of rhythmic movement of these mutant or restore to the rhythmic movement of leaves.This result fully indicates that there is a direct or indirect governing relationship between the GABA metabolism and the circadian clock.In summary,the exogenous GABA and its endogenous/metabolic mutants mainly affect the amplitude of the rhythmic movement of leaves.The results of this study provide the evidence that the intrinsic metabolic balance of GABA in plants is the prerequisite for maintaining the circadian rhythm.Further study of the regulation of GABA metabolism on circadian rhythms is expected to become a potential way to increase crop yield and breeding.