Natural variation, functional divergence, and local adaptation of nucleotide binding site sequences in Rhododendron (Ericaceae)

To further understand natural variation and local adaptation in the evolution of plant defense, we analyzed polymorphism data of nucleotide-binding site (NBS) sequences of Rhododendron at both the species and population levels. Multiple duplication events were found in NBS sequence evolution in Rhododendron genomes, which resulted in six clades: A-F. Our results of several NBS clade pair comparisons showed significant evolutionary rate changes based on differences in substitution rates between NBS-encoding protein clades (type I functional divergence). Pairwise comparisons of NBS clades further revealed that many amino acids displayed radical biochemical property changes causing a shift in amino acid preferences between NBS-encoding protein clades (type II functional divergence). Such divergent evolution of NBSs is likely a consequence of positive selection related to differentiation of recognition signals in response to different pathogens. Primers specific to clades B and C, which differed in the number of radical amino acid changes causing type II functional divergence and levels of nucleotide diversities, were further used to amplify population clades B and C NBS sequences of Rhododendron formosanum populations. Higher levels of net nucleotide divergences (measured by D _a) between R. formosanum populations were found based on NBS sequences of population clade B compared to population clade C, suggesting local adaptation of population clade B NBS sequences. Local adaptation can be further inferred for R. formosanum population clade B NBS sequences because of significant Φ _ST based on variation in nonsynonymous substitutions. Furthermore, local adaptation was also suggested by no significant correlation of population pairwise F _ST between population clades B and C in R. formosanum.