The Study Of Arabidopsis Transcription Factor MS1 Regulating Pollen Surface Protein Expression

Pollen coat is an important part of pollen surface material and eposits on the outer wall of pollen.It can avoid excessive drying pollen and protect pollen from ultraviolet radiation and pathogen invasion.In addition,pollen coat also plays a key role in pollen-stigma interaction.MS188 encodes a R2R3-MYB family transcription factor(TF)which directly regulates the sexine formation.MS1 is the first reported male sterile gene in Arabidopsis thaliana.MS1 encodes a protein with a PHD-finger motif.In this paper,we studied the regulation of MS188 and MS1 to pollen coat protein genes(PCPs).By comparing ms188 mutant with ms1 mutant,sexine layer was missing on the pollen surface of ms188 anthers,however the sexine in ms1 still existed but the pollen coat was completely absent.The expression of MS1 was barely detectable in ms188 mutants,indicating that MS1 acts downstream of ms188.Chromatin immunoprecipitation(ChIP)indicates that MS188 directly binds to the MS1 promoter in vivo.Electrophoretic Mobility Shift Assays(EMSA)confirms that MS188 directly binds to the specific site of the MS1 promoter in vitro.Dual-luciferase transactivation assay(Dual-LUC)indicates MS188 directly activates MS1 expression.pPCPs::PCPs-GFP constructs were generated and transformed into wild-type plants.GFP signals of all transgenic lines were initially observed in tapetum and transported into locule following tapetum degeneration.The expression of most PCPs was decreased in both ms1 and ms188,suggesting that these PCPs act downstream of MS1.ChIP indicates that MS1 directly binds to the GRP14 and GRP19 promoter in vivo.EMSA confirms that MS1 directly binds to the specific site of the GRP14 and GRP19 promoter in vitro.The transient activation assay showed that MS1 could directly activate the expression of GRP14 and GRP19 in the mesophyll cells of tobacco leaves.pMS188::MS1 construct was generated and transformed into ms188/+ heterozygote mutant.In the transgenic plants with ms188 homozygote background,the normal expression of MS1 in ms188 can restore the expression of PCPs,suggesting that MS1 is sufficient to activate partially PCPs expression in tapetum.These results reveal that in(MS188?MS1?PCPs)pathway,MS188 is a direct regulator for sexine formation and MS1 direct activate the expression of most PCPs for pollen coat formation during pollen development of Arabidopsis thaliana.