Molecular Mechanism Of Arabidopsis HXK1-CLF Complex In Glucose-mediated Gene Expression

The physiological process of sugar production through photosynthesis is essential to maintain the survival and development in plants.In addition to being a basic molecule of cells and a central molecule of energy metabolism,glucose is also a hormone-like substance that can be used as a signaling molecule to regulate the physiological functions,metabolism and gene expression of organisms,and glucose signaling is the oldest and most conserved signaling compared with other sugar signaling.However,how do plants recognize and transfer the sugar signals generated by the external and internal environments to regulate plant growth and development? A large amount of research is currently based on HXK1 that is the most important glucose-specific sensor in plants.Existing research shows that it can recruit some transcription factors to form a complex,thereby regulating the expression levels of the downstream target genes and finally regulating the entire process of plant growth and development.However,the role of epigenetic regulation in this process is currently unknown,even though there are already many reports about epigenetic changes induced by abiotic stresses.Among the epigenetic regulatory elements,Polycomb group(Pc G)and Trithorax group(Trx G)are the two most conserved protein families.Pc G protein which mainly exists in the form of two complexes of PRC1 and PRC2 is generally related to the inhibition of gene expressions.H3K27me3 is a marker that Pc G protein inhibits the transcription of target genes.Several members of the reported PRC2 and PRC1 complexes include three H3K27me3 methyltransferases: MEA,CLF,and SWN.The Trx G protein is generally associated with the activated expressions of genes which are usually H3K4 methyltransferase and ATP-dependent chromatin remodeling enzyme.Previous studies have shown that these two protein families usually exercise antagonism and eventually form a balanced and optimized state of gene expressions.Our laboratory previously found that the mutant of ATX5(atx5)that belonged to the Trx G protein family had a significant sugar-sensitive phenotype.Therefore,this study mainly used the mea,clf,and hxk1 loss-of-function mutants as experimental materials to explore whether the epigenetic element of Pc G protein family could participate in the HXK1-dependent glucose signaling pathway by using the methods of genetics,molecular biology,bioinformatics,and biochemistry.The experimental results were as follows:(1)The observation phenotypes of clf,swn and mea mutants under high glucose conditions demonstrated that the clf and swn mutants showed a glucose-insensitive phenotype.(2)The preliminary RNA-seq analysis of Col-1,clf,swn,and hxk1 materials treated with liquid Murashige and Skoog(MS)medium containing 6% glucose for 0 h and 1 h showed that CLF,SWN and HXK1 might co-regulate some targets together.(3)Through co-transforming the HXK1-GFP vector and the NLS-RFP vector with nuclear localization signal into Arabidopsis protoplasts,we observed At HXK1 was localized in the nucleus by using the fluorescence microscope.(4)Through Co-IP experiments,we found that At HXK1 and At CLF could interact with each other in vivo.(5)Through the observation of the phenotypes of clf hxk1 double mutant plants under high glucose conditions,we found that the clf hxk1 double mutant also showed a glucose-insensitive phenotype.Based on the abovementioned results,we suggest that At CLF can participate in the HXK1-dependent glucose signaling pathway to regulate the expression of the downstream target genes of HXK1 by forming a HXK1-CLF complex with HXK1.