Evolutionary Origins and Functional Diversification of 2′-O-Methyltransferases: Insights from Phylogenetic and Structural Analysis

Ribose 2'-O-methylation (Nm), a key RNA modification, is catalyzed by diverse 2'-O-methyltransferases (2'-O-MTases), yet the evolutionary trajectories of these enzymes remain poorly studied. Here, with a comprehensive collection of functionally validated 2'-O-MTases, we classified them into 11 families based on the distinct methyltransferase (MTase) domains. Homology searches across 198 species identified 6746 proteins, revealing the widespread distribution of 2'-O-MTases across the Tree of Life. Eight MTase domains (e.g., FtsJ, SpoU-methylase) existed both in eukaryotes and prokaryotes, indicating their ancient origin in the Last Universal Common Ancestor (LUCA). In contrast, the AdoMet-MTase, TRM13, and Trm56 domains are lineage-specific. Copy number expansion of most 2'-O-MTase families occurred as life evolved from prokaryotes to eukaryotes, where they might engage in more complex regulation of cell differentiation and development. Domain composition, Ka/Ks ratio, and domain structural analyses showed that purifying selection conserved catalytic domains across most families, despite the frequent integration of auxiliary domains. Notably, the FtsJ family diverged into three deeply separated lineages via remodeling the catalytic pocket, with each lineage specializing in the methylation of mRNA caps, rRNA, or tRNA. These findings illuminate the evolutionary trajectory of 2'-O-MTases, highlighting their ancient multiple origins and functional diversification.