Transcriptome Analysis Of Zar1 And Screening Ligands Of ZAR1 In Arabidopsis Thaliana

The molecular mechanisms of double-fertilization,zygote activation after fertilization,embryo development are key topics in plant reproductive development.Zygote asymmetrical division plays crucial role in pattern formation during early embryogenesis.Zygotic arrest 1(ZAR1)Proteins,belongs to the leucine-rich repeat receptor-like protein kinases family in Arabidopsis,which regulates the division of zygote and the cell fate of its daughter cells.ZAR1 is specifically expressed in central cell,egg cell and synergids in the mature embryo sac;after fertilization,ZAR1 is accumulated in zygote and endosperm.ZAR1 can interact with the G? protein and Ca M1,controlling the zygotic asymmetric division and the cell fate of its daughters.To explore the ligand of ZAR1-mediated signaling networks,we sequenced and analyzed m RNA of wild-type Ler and zar1-2,from the beginning of megasporogenesis to the end of zygotic asymmetric division in Arabidopsis thaliana.We found fifteen secreted small peptides,which belongs to CRPs(cysteine-rich peptides)family,were significantly different from ZAR1 or co-expressed with ZAR1,suggesting that these peptides may include the ligands of ZAR1.By yeast two-hybrid assays,we preliminarily identified that ZAR1 might interact with small peptides.ZI9 interacted weakly with the extracellular domain of ZAR1.ZI1,ZI10,and ZI13 had self-activation activity,their interactions with ZAR1 needs further confirmation.ZI2,ZI6,ZI14 interacted with ZAR1-LRR,whether these three small peptides are the ligand,it will be checked in pull-down with ZAR1 in vivo,expression pattern detection,their mutant phenotype analysis,to identify the true ligand of ZAR1 in regulating the asymmetric division of zygote.