| H2O2 production in plants is a hallmark of successful recognition of pathogen infection and pathogen-associated molecular patterns(PAMPs).Production of H2O2 is typically in apoplastic but is associated with intracellular immunity pathways that regulate disease resistance,such as systemic acquired resistance(SAR)and PAMP-triggered immunity(PTI).Our laboratory has proved that a plant aquaporin(AQP),plasma membrane intrinsic protein AtPIP1;4,acts to break the cytological obstruct between H2O2 production and functional performance.This role of AtPIP1;4 is essentially required for cytoplasmic transport of apoplastic H2O2 induced by a bacterial pathogen and two distinct PAMPs.In consequence,cytoplasmic H2O2 quantities substantially increase while SAR and PTI are activated to repress the bacterial pathogenicity,suggesting the apo-cytoplastic signal transduction affects immunity pathways.However,there are 13 AtPIPs in Arabidopsis,whether exists other AtPIPs can transport H2O2 besides AtPIP 1;4.AQPs can not only mediate the rapid and efficient transmembrane transport of water molecules,but also permeate H2O2,and participate in intracellular immune signal transduction by mediating the transport of intracellular cytoplasmic H2O2.With the sensitivity of yeast cells heterologously expressing AtPIPs to H2O2 was analyzed,and the ability of AtPIPs to permeate H2O2 was detected by using ROS probe DCF and special probe AR(Amplex Red)to judge the entry of H2O2 into the yeast cells.We found that yeast cells heterologously expressing AtPIP2;4,AtPIP2;5 and AtPIP2;7 had lower survival rate than yeast cells transfected with other AtPIPs,and had stronger ability to permeate H2O2 with stronger fluorescence intensity than those transformed into empty carriers.And we analyzed the induced expression of 13 AtPIPs in Arabidopsis thaliana inoculated with Pst DC3000(Pseudomonas syringae pv.tomato DC3000).It demonstrated that the AtPIP2;4,AtPIP2;5 and AtPIP2;7 were up-regulated significantly,suggesting that they might be involved in the resistance of Arabidopsis thaliana to pathogenic bacteria.Based on this,the three primers method was used to screen atpip2;4,atpip2;5 and atpip2;7 mutants,and we obtained three mutant homozygous lines.And then the overexpressing and complementing homozygous lines of AtPIP2;4,AtPIP2;5 and AtPIP2;7 were obtained by Agrobacterium-mediated transformation.Under biological or abiotic stress,plants will induce ROS production to regulate the defense and disease resistance response in the form of signaling molecules.H2O2 is not only the most important ROS signal and but also a Liaison factor for the immune response of internal and external plant cells.When plants are infected by pathogens or PAMPs,they would activate NADPH oxidase on the plasma membrane of plant cells and form H2O2 through a series of redox reactions.The resulting H2O2 is initially distributed in the apoplast,transported to the intracellular phase via AQPs on the plasma membrane,and subsequently involved in intracellular signaling,thereby increasing plant disease resistance.In this study,the resistance of AtPIP2;4,AtPIP2;5 and AtPIP2;7 to Pst DC3000 in Arabidopsis thaliana was studied by means of symptoms,bacterial content and callose deposition after inoculation with Pst DC3000 Overexpression of AtPIP2;4,AtPIP2;5 and AtPIP2;7 in Arabidopsis thaliana are used as experimental materials,after dyed by H2DCF-DA,AR and AUR,their ability to transport H2O2 are finally detected by laser confocal microscopy and multifunctional enzyme marker.The results showed that AtPIP2;4,AtPIP2;5 and AtPIP2;7 overexpressed strains were more resistant than wild type,and the mutants were more susceptible than wild type.It also mediates the pathogen-induced H2O2 to permeate the cell membrane and accumulate in the cell.Rice infection compels OsPIP 1;3 to switch its function,allowing PthXol to induce virulence by activating its target gene in an OsPIP1;3-dependent manner. |
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