Dynamic Distribution Of H₂O₂and The Gene Expression Analysis Of Its Metabolism-related Enzymes During The Interaction Between Wheat And Leaf Rust

Preliminary work in our laboratory have indicated that incompatibility combinationwheat （Triticum aestivum.） and leaf rust （puccinia triticina） presented a bimodal curvethrough detection H₂O₂generation changes with fluorescent probes DCFHDA, proving theH₂O₂burs of the programmed death of the host cell by pharmacological tests. To furtherexplore the dynamic distribution of H₂O₂and its relationship with HR, wheat （Triticumaestivum L.） varieties Lovering10（L10） respectively with wheat leaf rust physiological races165and260compose compatible and incompatible combinations and use DAB、CeCl3,specific marker of H₂O₂respectively, by means of optical microscopy and transmissionelectron microscopy to observe the Dynamic distribution of H₂O₂in the infection process ofwheat leaf rust. Then we analysis H₂O₂related gene expression by RT-PCR technologyanalysis to explore molecular mechanism of H₂O₂in transcriptional level. The main resultsare as follows:1. Dynamic distribution characteristics of H₂O₂during the interaction between wheatand leaf rust in the histological level with optical microscope. The test results showed that theH₂O₂distribution in compatible and incompatible combinations had no difference between0hand12h and H₂O₂mainly concentrated on the guard cells of infection point. In theincompatible combination at24h after infection, the mesophyll cell infected distributed asmall amount of H₂O₂.48h cells slightly distorted were full with distribution of a largenumber of H₂O₂.72h the mesophyll cells collapsed were distributed with reducedH₂O₂.Compatible combination was always not observed H₂O₂staining at late inoculation.2. Dynamic distribution characteristics of H₂O₂during the interaction between wheatand leaf rust in the subcellsular level with electron microscopy techniques. The test resultsshowed that in the compatible and incompatible combinations4h after the interaction theguard cell wall and mesophyll cell wall below the appressorium had H₂O₂distribution. Andthe amount of H₂O₂in the incompatible combination4h was more than that in the compatiblecombination. In the incompatible combination,8h began to form substomatal cysts andmesophyll cell within a lot of H₂O₂distribution. On the contrary, in the compatiblecombination we couldn’t detect any H₂O₂above area in the incompatible combination. At12h there was no difference between compatible and incompatible combinations about thedistribution of H₂O₂.Both weren’t detected H₂O₂. Between24h and72h in the incompatiblecombination, we observed a large number of H₂O₂in haustorial mother cell、haustorium、abortive haustorium、 the interaction of guard cell wall and mesophyll cell wall and healthycells. On the contrary, in the compatible combination we couldn’t detect any H₂O₂infungus and host cells at the same time. 3. RT-PCR results showed that in the incompatible combination, high expression ofTaSOD1and low expression of TaCTAA、TaAPX1together led to the imbalance thatproduction and elimination of H₂O₂, then accumulated a large number of H₂O₂.In thecompatible combination, high expression of TaSOD1, TaCTAA and TaAPX1maintain balancethat production and elimination of H₂O₂, then lost Oxygen burst.Based on the above results, we initially speculate that wheat with leaf rust interactionH₂O₂is mainly derived from host cell membrane and host cell wall; early H₂O₂play a role ofsignal that lead to defense responses produced by host cells, later period H₂O₂directly takepart in bactericidal action and promote HR produced by host cells. Also later period H₂O₂play a role of signal that lead to defense responses produced by peripheral cell around HR.