Mechanism Study Of Brassinolide And Sugar Signals Crosstalk To Regulate The Maintenance And Differentiation Of Plant Stem Cells

Plant stem cells similar to animal stem cells,regulate the morphogenesis of living individuals.But the difference is that plants adopt a post-embryonic morphogenesis model to regulate the occurrence,development,dormancy and aging of their organs to adapt to the environment by sensing the external environment.by sensing the external environmental signaling.Therefore the function of stem cells is more direct and important for plant.According to development process,plant stem cells include the apical stem cells in embryonic stage,the lateral stem cells in late embryos,and the regenerative stem cells established by the transformation of somatic cell fate during regeneration.These different types of stem cells determine the variety of plant morphogenesis in nature and their adaptability to the environment.Therefore,in order to further study the molecular mechanism of plant stem cell fate determination,this paper focuses on Brassinolide(BR)signaling transcription factor BRASSINAZOLE-RESISTANT1(BZR1)and the key kinase Target of Rapamycin(TOR)of Sugar signaling.Two types of root organs were studied,which are the main roots originating from the apical meristem(embryo formation)and the adventitious roots derived from regenerative stem cell.Furthermore,some molecular mechanisms for regulating the fate of stem cells have been discovered.The experimental results include the following two aspects:1,Sugar signaling and Brassinolide synergistically regulate apical quiescent center(QC)cell division:The study found that sucrose promotes QC division.Under sucrose-free conditions,the QC division rate was 4%,while the QC division rate increased significantly at 1%,3%and 5%sucrose concentrations,followed by 35%,48%and 60%.The same concentration of non-metabolizing sugar had no effect on the QC division rate.It is indicated that the regulation of sugar molecules involved in QC stem cell division activity is not caused by osmosis,but is related to sugar signaling pathway.For further,It was found that the QC division rate of the tor mutant was reduced by about 50%,and the QC division rate of the HXK1 kinase mutant gin2-1 was only decreased by about 20%,indicating that sugar promotes QC division requiring TOR kinase and HXK1,and TOR kinase is more important.It was found that BR treatment can restore tor QC division phenotype,and BR-enhanced mutant bzrl-1D can also restore tor mutant less QC division rate phenotype though tor/bzrl-1D double mutant phenotype analysis.Besides,the BR receptor deletion mutant bril-301 was found to be insensitive to sugar,while bzrl-1D was hypersensitive to sugar,confirming that sugar signaling regulates QC stem cell activity depending on BR signaling.At the molecular level,it was found that sugar can inhibit the degradation of BZR1 protein in root tip stem cells.Treatment with exogenous TOR kinase inhibitor AZD8055 was showed that the expression level of WOX5 decreased,indicating the kinase activity of TOR can promote the expression of WOX5.Under the same treatment conditions,the expression level of WOX5 in bzr1-1D is higher than that of wild type WOX5.This experiment clarified that BZR1-D protein promote WOX5 expression because it is located in the nucleus without being degraded,though the autophagy pathway was stimulated during TOR kinase was bloched by AZD8055 treatment.It is proved that the sugar signaling inhibits the degradation of BZR1 protein through the TOR kinase,subsequently increases the expression of WOX5 in QC cells and promotes QC division.This study elucidated that sugar as a signaling molecule promotes the division of apical stem depending on BR signaling,providing new clues for the study of apical stem cell activity.2,Both sugar signaling and Brassinolide regulate adventitious root formation:Adventitious root is the root organ formed by the division and differentiation of the original layer cells with stem cell activity in the body after the plant is injured in vitro.Studies showed that sugar signaling promotes adventitious root formation requiring TOR and HXK1;while high concentration of BL and BR downstream transcription factor BZR1 significantly delayed and inhibited adventitious root formation.At the molecular level,BZR1 was found to inhibit the expression of WOX11 and WOX12.It was observed that 35S::WOX11 partially restored the phenotype of bzr1-1D with less adventitious root formation by genetic analysis,indicating that BZR1 was located upstream of WOX11.By SEM scanning electron microscopy and qRT-PCR analysis of WIND family genes,it is believed that the white cell mass at the bzr1-1D wound may not be callus.However,whether the BR and sugar signaling are coordinated with each other and participate in the formation of adventitious roots remains to be further studied.This study was carried out in vitro,indicating that the BZR1 transcription factor,as a negative regulator,is involved in in vitro regeneration.