Function Of ABIP1 Involving In Pib-AvrPib Interactions In The Rice Blast Pathosystem

Rice blast, caused by ascomycete fungus Magnaporthe oryzae, is a destructive and recurrent problem in all rice-growing regions of the world. The use of resistance genes in crop breeding programs has been, and will undoubtedly remain the major means for disease control,and it is the most cost-effective and environmentally friendly strategy. The interactions between the rice and Magnaporthe oryzae is in line with the gene-for-gene hypothesis, the in-depth study of the interactions between the avirlence gene and the resistance gene may provide a better understanding of host-pathogen pathosystem and have insights into control blast disease.In the previous work, we have isolated the avirulence gene AvrPib by map-based cloning, and set about studying of the AvrPib interacting protein 1(ABIP1). In order to illustrate the function of ABIP1 involving in Pib-AvrPib interactions in the rice blast pathosystem, the main contents and results are as follow: 1. The fuctions of Pib, AvrPib and ABIP1In the previous work, we have known the subcellular localization of Pib, AvrPib and ABIP1. We use Agrobacterium-mediated transient expression system in Nicotiana benthamiana to futher analyze the fuctions of Pib, AvrPib and ABIP1. And it showed the HR-like response was induced only by PibCC. After fusing with nuclear localization signal(NLS) or nuclear export signal(NES), the HR-like response of PibCC was disappeared. The results showed that the HR-like response was induced only when the subcellular localization of PibCC fusion proteins were nucleocytoplasmic. 2. Interactions among Pib, AvrPib and ABIP1It has been proved that the non-signal peptide of AvrPib(AvrPib23-74) have interaction with PibCC and PibNBS1 in the previous work. In order to better understanding of the potential functions of the signal peptide(AvrPib1-22), a bimolecular fluorescence complementation(Bi FC) assay was applied to confirm the interactions between AvrPib1-22. The results showed that AvrPib1-22 was only interacted with PibNBS1. While the Pull-down assay showed that AvrPib1-22 have no interactions with PibCC and PibNBS1.In order to illustrate the molecular mechanisms of ABIP1 in the interaction model of AvrPib and Pib, the Bi FC system was applied to confirm the interactions among ABIP1, AvrPib and Pib. No interactions were found between ABIP1 and Pib, and these results were verified via Agrobacterium-mediated transient expression system.However, AvrPibFL and AvrPib23-74 could interct with ABIP168-172 and ABIP168-278 in both the cytoplasm and the nucleus; but the Pull-down assay unexpectedly have different results that ABIP1 and AvrPib have no interactions.The interactions among Pib, AvrPib and ABIP1 was further verified via Agrobacterium-mediated transient expression system. Comparing with the control group of PibCC, the combination of PibCC, AvrPib and ABIP1 may suppressed the auto-activation of PibCC. 3. Interactions between the phosphorylate mutations of ABIP1 and AvrPibTo further elucidate the molecular mechanism of ABIP1, we have observed the subcellular localization of the phosphorylate mutations(ABIP168-172 S139 A, ABIP1 FL S103A and ABIP1 FL S139A). The fusion proteins showed that the subcellular localization of the mutations have not differernce with ABIP168-172 and ABIP1 FL. And then, the Y2 H assay was applied to confirm the interactions between AvrPib and the mutations of ABIP1, the result showed only the locus of 103 and 139 have influenced the interactions between ABIP1 FL and AvrPib1-22; while there were no influence on the interactions between ABIP1 and AvrPib1-22 in the Pull-down assay, and the result of Bi FC system have confirmed this.