Evolutionary fates of gene-body methylation and its divergent association with gene expression in pigeonpea.

Pigeonpea (Cajanus cajan L. Huth) is an agronomically important legume cultivated worldwide. In this study, we extensively analyzed gene-body methylation (GbM) patterns in pigeonpea. We found a bimodal distribution of CG and CHG methylation patterns. GbM features- slow evolution rate and increased length remained conserved. Genes with moderate CG body methylation showed highest expression where as highly-methylated genes showed lowest expression. Transposable element (TE)-related genes were methylated in multiple contexts and hence classified as C-methylated genes. A low expression among C-methylated genes was associated with transposons insertion in gene-body and upstream regulatory regions. The CG methylation patterns were found to be conserved in orthologs compared with non-CG methylation. By comparing methylation patterns between differentially methylated regions (DMRs) of the three genotypes, we found that variably methylated marks are less likely to target evolutionary conserved sequences. Finally, our analysis showed enrichment of nitrogen-related genes in GbM orthologs of legumes, which could be promising candidates for generating epialleles for crop improvement.