Receptor-ligand Pair Regulate Pollen Hydration In Arabidopsis Thaliana

The interaction between male and female gametophytes in angiosperms is mainly composed of pollen adhesion,pollen hydration,pollen germination,pollen tube guidance to the female gametophyte,and pollen tube reception.Studies have mainly focused on the pollen tube guidance to the female gametophyte and the pollen tube reception,however little is known about the signaling pathway of pollen-stigma interaction.In this project,we found that ANJEA and FERONIA receptor kinase highly expressed in the stigma of Arabidopsis thaliana.Through genetics,molecular biology,cell biology,and biochemical approaches,we identified the key components of pollen-stigma interaction and explored the mode of ligand-receptor interaction in the process of affinity pollination.Staining of p ANJ::GUS and p FER::GUS transgenic materials revealed that both ANJEA and FERONIA were expressed in stigmatic papilla cells.Observations of p ANJ::ANJ-GFP and p FER::FER-GFP transgenic plants showed that both ANJEA and FERONIA were located in the plasma membrane of stigmatic papilla cells.Further phenotypic analysis of the single and double mutants of anj and fer revealed that the stigma of the mutant defective in pollen adhesion and pollen hydration rate.Subsequent pollen tube growth rate of the mutant was faster than that of the wild type.These results indicated that stigma highly expressed ANJEA and FERONIA participated in pollen-stigma interaction.Bi FC and pull-down assays confirmed the interaction between ANJEA and FERONIA through the extracellular domain,indicating that the two can form a protein complex to participate in the signal pathway of pollen-stigma mutual recognition.Multiple members of RALF family in Arabidopsis have been found to participate in a variety of signaling pathways as ligands of members of Cr RLK1 L family in Arabidopsis.In this project,the interaction between pollen and stigma highly expressed RALFs and ANJEA/FERONIA was confirmed by bioinformatics analysis and interaction experiments.Further construction of related ralfs mutants and preliminary analysis of pollen hydration related phenotypes revealed that RALF33 secreted by stigma and RALFs highly expressed in pollen may mediate pollen-stigma interaction as ANJEA/FERONIA ligand during pollination.In summary,this study identified that the RALF small peptide-ANJEA/FERONIA receptor kinase combination regulated pollen-stigma interaction and expanded the understanding of stigma affinity response mechanism.It provided an important theoretical basis for elucidating the mechanism of pollen-stigma interaction.