Structure And Phylogenetic Analysis Of Three Classes Of Rice Disease Resistance Homologues

Plants have formed their own defense systems in the long history of coevolution with various pathogen, in which disease resistance genes (R genes) play important roles. Since the first R gene Hml has been cloned in 1992, more than 40 have been cloned from many plant species, which includes rice, tomato, and so on. Rice is a most important crop in the world, from which, some R genes, such as Xa21, have been obtained. It can strong the cultivated rice species when R genes were introduced to them, but one or two R genes is not sufficient to meet the the goal in plan. So far, we have obtained the whole genome sequences of two subspecies (Indica and Japonica) in rice, which were finished in the strategy of whole genome shotgun. This gives us a intention to obtain the whole resource of rice disease resistance homologues using bioinformatics, which can prepare for the R gene cloning and seed improvement in next step.1. Distribution and structure analysis of rice LRR-TM homologuesLRR-TM (Leucine-rich repeat plus transmembrane) domains are an importantcomponent with an extra-cellular LRR domain and a coil structure spanning the cell membrane in plant disease resistance genes. Through the homology search, we have collected all 174 and 149 candidates for homologues from indica and japonica genome respectively. These candidates exist in clusters with a large part, and singles on the 12 chromosomes in each genome. In theses candidates, 14 from each genome have the specific structure with both transmembrane subdomain and signal peptide sequences, which were regarded as the LRR-TM homologues according to their 4 coutparts in tomato have the same characters. From the clusters distribution, we presume that most deletions and/or amplifications are occurred there. Additionally, we compared the similarities between the sequences from two subspecies, and found most (65% from each genome) have high similar countparts in the other, from which, we think that most LRR-TM candidates are formed before the divergence of the two subspecies, and other 35% are amplified after that point. From the phylogenetic analysis of all LRR-TM analogues, two LRR-TM analogues from indica, five from japonica and four Cf genes from tomato locate in the same clade. which may suggest that they evolve from a common ancestor. The distribution in genome, structure and phylogenetic information of rice LRR-TM candidates will give important clue for identifying the rice disease-resistance genes, as will be shown later.2. Distribution and structure analysis of rice STK homologuesSTK (serine-threonine kinases) is an important form in plant disease resistance genes that composed by a single function Serine/Threonine kinase domain. Using the protein sequences of members from STK R genes family as queries, genome-wide searches for the STK R gene homologues are made in both subspecies of rice (Indica and Japonica), we have found 142 and 132 homologues with STK domain respectively, most of which have no transmembrane subdomain and signal peptide, which is similar to Pto in tomato. These homologues distribute as either single gene or gene cluster in the genomes, contain eight or nine conserved sequences with 20 to 57 amino acids respectively, and more than half of these STK homologues in each genome have one or more high similar (>80%) counterparts in the other. From the phylogeny tree constructed by STK related sequences from rice, Arabidopsis, tomato and so on, we could find non-rice sequences dispersed among rice ones, and sequences from tomato are clusterd in a few branches. These findings may lead us to assume that most of rice STK homologues formed before the divergence of two rice subspecies.3. Distribution and structure analysis of rice NBS-LRR homologuesNBS-LRR homologues, which is common in disease resistance genes, contains NBS(Nucleotide binding site) and LRR (Leucine rich repeat). We collected 18 sequences from plant NBS-LRR family, which were used as queries to do genome-wide searches in both subspecies of rice (Indica and Japonica), and derivated 281 and 275 NBS-LRR homologues respectively. Most of these homologues have no transmembrane subdomain and signal peptide, which is similar to most of the cloned NBS-LRR R genes in plants. These homologues distribute as either single gene or gene cluster in the genomes, similar to LRR-TM and STK homologues, contain 10 conserved motifs with 20 to 51 amino acids in each genome respectively. In each genome, more than half of these NBS-LRR homologues involved in the high similar (>80%) sequences pairs. A phylogeny tree constructed by NBS-LRR related sequences from rice, Arabidopsis, tomato and so on, we could find non-rice sequences are dusted together, which shows a long distance between rice homologues and non-rice ones.4. Comparison of codon bias and phylogentic analysis between homologues in the three classesA further detail study on the nucleotide level in the three classes diseases resistance(R) homologues in Indica and Japonica, shows various base composition on each site of codons, and different usage for synonymous codons. Comparison for R sequences from rice and tomato told us a great variety between the two. We checked of theconserved domains in rice R homologues identified before with different E values once more, and those identified homologues varies much, which may be the result of the long and complicated evolution. Assume that the rate of synonymous substitution are constant in both tomato and rice with a value of 6.5 X 10" , we predict that the evolution divergent time between rice and tomota homologues is from 240 to 540 million years ago (Mya), and most rice homologues formed before the divergence between its two supspecies (Indica and Japonica).