Functional Study Of Gibberellins Effects On The Initiation Of Axillary Meristem In Arabidopsis

As sessile organisms,plants need to constantly adapt their growth and development to the changing environment by adjusting the occurrence,development,dormancy,and aging of their organs.The remarkable plasticity of plant development is believed to rely on networks of interconnected signal transduction pathways that integrate multiple hormonal and environmental signals coordinately regulating the characteristics of plant meristems.Plant shoot branching pattern affects plant architecture and is pivotal for developmental plasticity and crop yield.Shoot branches develop from axillary meristems(AMs)that initiated in the boundary zone between leaf primordia and the shoot apex,which is regulated by multiple endogenous phytohormonal signals and different environmental cues.Gibberellin is an important phytohormone in plants and regulates many developmental processes in plants,including seed germination,cell elongation,stomatal development,and photomorphogenesis.Here,in this study,we had combined the advantage of plant physiology,genetics,molecular biology and biochemistry to gain a deep insight into the molecular mechanism of gibberellin regulation of the initiation and differentiation of axillary meristem in Arabidopsis1.Gibberellin represses the initiaion of axillary meristem.Whether the spraying of gibberellin in vitro or the deficiency of DELLAs protein which are the key repressors of gibberellin signalling,result in the reduction of plant lateral shoot formation.On the contrary,the use of gibberellin biosynthesis inhibitor PAC to treat plants and the gibberellin deficent mutants gal-3 all show an increased phenotype of axillary buds,indicating that gibberellins inhibit the initiation of axillary meristem,which in turn lead to a decrease in plant lateral buds.2.DELLAs protein interact with SPL9 in vivo and in vitro.To analyze the molecular mechanism that gibberellin regulates the initiation of axillary meristem,we screened and analyzed the interaction between DELLAs protein RGA and transcription factors that specifically expressed in axillary meristem.In this paper we did a deep research on one of the transcription factors SPL9(SOUAMOSA PROMOTER BINDING-LIKE 9).At first,we used yeast two-hybrid,bimolecular fluorescence complementation and protein pull-down assay to analyze the interaction between RGA and SPL9.The experimental results show that RGA interacte with SPL9 both in vivo and in vitro.3.miR156-SPL9 regulates the formation of lateral buds in plants.SPL9 is a SBP family transcription factor regulated by miR156.The miR156 over-expression plants resulted in the increase of plant lateral buds,while the rSPL9 plants that the expression of SPL9 would't be degraded by miR156 significantly inhibited the formation of plant lateral buds.These results indicate that the miR156-SPL9 pathway plays an important regulatory role in the lateral bud formation of plants.4.The association of DELLAs to SPL9 inhibits the transcriptional regulation of SPL9 to LAS,the key factor in the initiation of lateral buds.Our experimental results show that SPL9 could directly bind to the promoter of LAS to inhibit the expression of LAS.The association of DELLAs inhibits the binding of SPL9 to the LAS promoter and further promotes the expression of LAS.5.LAS regulates the expression of the GA degradation gene GA2ox4.LAS,a key transcription factor regulating the initiation of lateral bud meristem,could directly bind to the promoter of GA2ox4 and promote the expression of GA2ox4.This results in the low concentration of GA in the initiative region of lateral buds,and in turn leads to enrichment of DELLAs and inhibition of the transcriptional activity of SPL9,then increases the transcript levels of LAS,forming a positive feedback loop that regulates the formation of lateral buds.Systematic analysis of the molecular mechanism of gibberellin regulation of axillary meristem initiation and differentiation not only helps us to understand the molecular mechanism of axillary meristem development,but also provides theoretical support and technical assistance for our next step to cultivate a new breed of high-yield and stress-tolerant with gibberellin,and so as to promote China's agricultural modernization and biotechnology industrialization as soon as possible.