Screening And Verifying Disease Resistance Related Proteins And Stomata Developmental Proteins That Interacted With Phosphatase TOPP4 In Arabidopsis Or SFC In Rice

This paper contains two parts: the first part verified the proteins that interact with Arabidopsis phosphatase TOPP4 in plant immune pathway using yeast two hybridization assays;the second part screened the proteins that interact with SFC which plays an important role in stomata development in rice using yeast c DNA library.Part 1: previous work of our laboratory has found that Arabidopsis phosphatase TOPP4 is involved in many biological processes,such as the regulation of GA signal pathway,auxin polar transport,and light signaling pathway.Recently,we find the phenotype of topp4-1 could be partially restored by the mutant of key regulators of the SAR pathway.This result suggests that TOPP4 plays a role in plant immune signaling pathway.This work is based on that find.The results of RT-PCR show that the expression of two CC type R proteins,RPS2 and PRS5,is up-regulated in topp4-1.The genetic phenotype of double mutants,rps2 topp4-1 and rps5 topp4-1,showed the TDNA insertion mutants of rps2 and rps5 fail to restore the phenotype of topp4-1.To explore the proteins that interacted with TOPP4 in plant immune pathway,we screened 12 key proteins which had been identified in immune signal pathway previously by using yeast two hybridization assays.The results show that TOPP4 could interact with MPKX,MKSY,MPK2,and EDS1.While besides binding these proteins,topp4-1 could interact with RIN4.The interactions between TOPP4,topp4-1 and MPKX,MKSY are further confirmed by Bi FC experiments,suggesting that TOPP4 may regulate plant immune through changing the phosphorylation of MPKX and MKSY.Part 2: F-box protein is a general kind of proteins with a conserved ‘F-box' motif at the N terminal.As a component of ubiquitin-proteasome,F-box protein could recognize substrates of this complex.F-box protein plays a role in various biological processes in plants.However,the revealing of its function in monocots is very rare until now.Previous studies in our lab find a mutant of F-box protein in rice,sfc,which show stomatal defect in subsidiary cells.To uncover the target proteins of the F-box protein that regulate stomatal development in rice,we have screened rice leaf primordium c DNA library using yeast two hybridization system.As a result,22 positive clones are selected through library screening.Analyzing these selected positive clones,we find that these genes are mainly involved in various aspects of plant growth and development,such as the stability of microfilaments and microtubule,cell division and cell cycle.Six candidate genes are chosen to confirm their interactions with SFC.Yeast two hybridization assays show that Os CK? and Os Rhmdb? could interact with SFC.Further study on the relationship between SFC and Os CK ? will uncover the important mechanism of stomatal development in monocots.