Rapid reorganization of resistance gene homologues in cereal genomes

Abstract

We used conserved domains in the major class (nucleotide binding site plus leucine-rich repeat) of dicot resistance (R) genes to isolate related gene fragments via PCR from the monocot species rice and barley. Peptide sequence comparison of dicot R genes and monocot R-like genes revealed shared motifs but provided no evidence for a monocot-specific signature. Mapping of these genes in rice and barley showed linkage to genetically characterized R genes and revealed the existence of mixed clusters, each harboring at least two highly dissimilar R-like genes. Diversity was detected intraspecifically with wide variation in copy number between varieties of a particular species. Interspecific analyses of R-like genes frequently revealed nonsyntenic map locations between the cereal species rice, barley, and foxtail millet although tight collinear gene order is a hallmark of monocot genomes. Our data suggest a dramatic rearrangement of R gene loci between related species and implies a different mechanism for nucleotide binding site plus leucine-rich repeat gene evolution compared with the rest of the monocot genome.

Figure 1

Phylogenetic tree of monocot R gene homologues and characterized dicot NBS-LRR genes. Amino acid sequences of rice (red) and barley (green) NBS-LRR genes were aligned by using the clustal w program (30). The phylogenetic tree was constructed by using the neighbor-joining method of Saitou and Nei (31). For comparative purposes characterized dicot NBS-LRR genes N (tobacco), M and L6 (flax), RPS2, Rpm1, and Rpp5 (Arabidopsis), and Prf and I2C-1 (tomato) have been included (1). Homologues or R genes sharing ≥80% DNA sequence similarity are marked by gray shaded circles.

Figure 2

Copy number variation of NBS-LRR genes in cultivars of rice and barley. Genomic Southern analysis for (A) the rice NBS-LRR probe r2 in six rice cultivars: Npb, Nipponbare; Kas, Kasalath; IR20; 6383; Ch45, Chugoku 45; and IR24. The number of bands range from two minor cross-hybridizing bands in Ch45 to 15 major bands in IR24. Analysis of segregants from the cross Ch45 × IR24 revealed that all NBS-LRR copies detected by r2 in Ch45 map to a single locus. (B) The barley probe b9 shows the diversity in copy number in six barley cultivars. Note that only two weakly cross-hybridizing signals are detected in cv. Franka but five prominent signals in cv. Igri.

Figure 3

Comparative mapping of R gene homologues in the monocot species rice, barley, and foxtail millet. A circle diagram according to Moore et al. (16, 17) was chosen to visualize syntenic relationships that align the genomes of barley (green), rice (red), and foxtail millet (blue). Map locations of NBS-LRR genes that could be mapped in at least two of the three tested species or are present in RHCs are given. Syntenic map positions are marked by bold red spokes and nonsyntenic R gene homologue loci are boxed in black. Clusters containing at least two highly divergent NBS-LRR genes in rice and foxtail millet (RHC-A to RHC-D) are highlighted in the periphery. Note that the rice NBS-LRR probe r2 detects loci in rice (Os-r2) and foxtail millet (Si-r2.3) that are organized in RHC-A and RHC-D (indicated by dotted black line). The barley NBS-LRR probe b6 detects two loci (Si-b6.1 and Si-b6.2) in foxtail millet that are also organized in different mixed clusters (RHC-D and RHC-E; indicated by a dotted black line). Positions of chromosome insertions are indicated by solid black lines; barley chromosomes are numbered 1H to 7H, rice chromosomes 1 to 12, and foxtail millet chromosomes I to IX. S and L denote the short and long arm of each chromosome.