Mechanism Study Of Brassinosteroid And SPL9 Crosstalk To Regulate The Leaf Age Development And Flowering In Arabidopsis

The growth,development and morphology of life in nature are closely related to age.Age is an irreversible process,and all living things undergo a transition from the juvenile to the adult phase,enter the reproductive phase and eventually to senescence and death.The effect of phase change on growth and development of vegetative stage is mainly reflected in leaf morphology in Arabidopsis.Previous studies showed that miR156 and its target gene SPLs are key regulators of plant age.Brassinosteroids,as sterol hormone,play important roles in all aspects of plant growth and development.The BR deletion mutant showed the longer vegetative period and early flowering phenotype.Does BR also participate in regulating the leaf age development and flowering of plant?Furthermore,does BR and miR156 crosstalk to regulate plant growth and development?To answer these questions,we use plant physiology,genetics,molecular biology,and biochemistry to study the molecular mechanism of BR regulating plant age and the molecular mechanisms of BR and SPL9 in regulating leaf morphological changes and flowering.Based on these experimental results,we made the following conclusions:1.BR regulates the leaf age development in Arabidopsis.By analyzing the phenotype of length/width and blade base angle of BR mutants in Arabidopsis,it was proved that BR can promote the age development of Arabidopsis leaf.RT-qPCR experiments prove that BR can promote the transcriptional expression of SPL,which may be the reason why BR can regulate leaf age development.2.BR is inolved in SPL9 regulating the leaf age development and flowering process in Arabidopsis.Through genetic analysis,we found that SPL9 can restore the phenotype of hypocotyl and leaf morphology of the BR deletion mutant det2,suggesting that BR plays an important role in SPL9 regulating the leaf age development and flowering in Arabidopsis.3.SPL9 regulates the age development of Arabidopsis leaves and flowering depend on BR.SPL9 can promote the elongation of mature leaves of Arabidopsis,the production of abaxial epidermal hairs and flowering.But the function of SPL9 is also inhibited when the BR signal weakens,which indicates that SPL9 regulates the age.4.SPL9 and BZR1 interact in vivo and in vitro.In order to further study the relationship between BR signaling and SPL9 transcription factors,we screened for proteins that interact with SPL9 in the BR signaling pathway.Through yeast two-hybrid experiments,protein pull-down experimentsand rBiFC technology,we found that SPL9 can interact with the presence of an important transcription factor BZR1 in the BR signaling pathway.5.SPL9 promotes cell elongation.Through the BR sensitivity experiment,we found that SPL9 gain-function plants were super sensitive to BR,but weakened to BR synthesis inhibitor PPZ.In addition,RT-qPCR experiments showed that SPL9 promoted the expression of key genes PRE1,PRE5,and EXP8 in cell elongation,indicating that SPL9 plays a key role in regulating cell elongation.In summary,we constructed a model for the regulation of age development and flowering of arabidopsis leaves by BR signal and SPL9.BR can promote the age of Arabidopsis and regulate the phenotype of leaves.BR and SPL9 are synergistic in regulating leaf age development and flowering in Arabidopsis.The interaction of transcription factors BZR1 and SPL9 in the BR signaling pathway can regulate leaf morphology by promoting cell elongation genes PRE1,PRES and EXP8,as well as jointly regulating the flowering process.