Evolution Mechanism Of Rice Bacterial Blight Resistance Gene Xa23 And Analysis Of Important Amino Acids Associated With HR Function Of XA23

Bacterial blight caused by Xanthomonas oryzae pv.oryzae?Xoo?is the most devastating bacterial disease in the world's major rice producing areas.The bacterial blight resistance gene Xa23 originated from Oryza rufipogon in China.The Xa23 confers wide-spectrum and high resistance to all the natural occurring Xoo races,and completely dominant,disease resistant throughout the growth period,with great value.Xa23 belongs to the Executor R gene.The EBE_AvrXa23vrXa23 located in Xa23 promoter can bind to AvrXa23,a TALE?Transcription activator-like effector?widely existing in Xoo strains,to induce the transcriptional expression of Xa23 gene.XA23 protein can induce HR in infected host cells,directly killing the infected cells to restrict the continued propagation of bacteria in the host and thus play a role in bacterial blight resistance.This paper includes two parts:the first part is to explore the evolutionary pathway and evolutionary mechanism of Xa23 gene;the second part is to detect and discover the amino acids/elements related to HR function in XA23 protein sequence as well as to analyze the properties of related amino acid residues.The main results obtained are as follows:Study on the evolutionary pathway and evolutionary mechanism of Xa23 gene:1.Xa23 alleles only appeared in the Oryza genus,and the coding region sequences of Xa23 alleles are highly conserved in Oryza species.2.There is no EBE_AvrXa23?28 bp?located in the promoter of xa23 in the natural cultivated rice by blasting the Rice SNP-Seek Database,indicating that there is no dominant Xa23 gene in the naturally-cultivated rice with known sequence.3.We collected 278 representative rice materials and only 97 rice lines contain Xa23/xa23 allele,suggesting that Xa23 alleles might be not widely distributed in Oryza genus.4.We found that the nucleotide sequence diversity of Xa23 alleles promoter is significantly higher than that of coding region.Therefore,based on the promoter sequence,Xa23 alleles could be recombined into five haplotypes?Pro-A/B/C/D/E?,of which Pro-A is the only haplotype containing EBE_AvrXa23.We inoculated these 97 rice lines with the Xoo strain PXO99^A and found that only the haplotype Pro-A confers resistance to bacterial blight disease.There is no significant difference in geographical distribution of the five Pro haplotypes.Pro-A exists only in wild rice,while Pro-B/C/D/E are distributed in both wild and cultivated rice,with the highest proportion of haplotype Pro-B?26.8%?.5.Xa23 originated from wide rice,and bottleneck effect may occur in Xa23 alleles during domestication and breeding of rice.Study on HR-related amino acids/elements in the XA23 protein sequence:6.All XA23 alleles proteins contain three transmembrane domains and show highest identity across the transmembrane domains.7.All the Xa23 alleles can cause HR,indicating that the sequences or elements related to HR function are conserved and complete in XA23 proteins.8.In XA23 sequence,the first 1-6 aa from the N-terminus and 1-9 aa from the C-terminus are HR-related essential amino acids,and the acid-base properties of the critical amino acids did not affect the HR phenotypes.Moreover,mutations in each transmembrane dormain can influence the HR phenotype,demonstrating that all the three transmembrane domains are important determinant of HR function.In conclusion,this study cloned the conserved Xa23 alleles in different rice species and discussed the origin and evolutionary mechanism of Xa23 gene.At the same time,we detected the HR-related amino acid residues in XA23.We speculate that these results are of great guiding significance for the discovery of more Executor R genes and for simplification of the Executor R protein structure in the future.