Evolutionary divergence of CXE gene family in green plants unveils PtoCXEs overexpression reduces fungal colonization in transgenic Populus

Plant enzymes significantly contribute to the rapidly diversified metabolic repertoire since the colonization of land by plants. Carboxylesterase (CXE) is just one of the ubiquitous, multi-functional, and ancient enzymes, that has particularly diversified during plant evolution. This study provided a status on the CXE landscape within Viridiplantae. A total of 784 CXEs were identified from the genome of 31 plant species representing 9 major lineages of sequenced Viridiplantae, and divided into five clades based on phylogenetic analysis. Clade I CXE genes may be of bacterial origin, and then expanded and diversified during plant evolution. Clade II was first gained in the ancestor of bryophytes after colonization of land by plants, Clade III and Clade IV in ferns which were considered the most advanced seedless vascular plants, while Clade V in seed plants. To date, the functions of CXE genes in woody plants remain unclear. In this study, 51 CXE genes were identified from the genome of Populus trichocarpa and further divided into eight classes. Tandem and segmental duplication events both contributed to the expansion of CXE genes in Populus. Although CXE genes were proven to enhance resistance to pathogens in many herbaceous species, relevant researches on forest trees are still needed. In this study, pathogen incubation assays showed that overexpressing of six Class VI CXEs in Populus tomentosa, to a greater or lesser degree, reduced colonization of detached leaves by fungus Cytospora chrysosperma. A significant difference was also found in functional divergence patterns for genes derived from different gene duplication events. Functional differentiation of duplicated CXE genes in Populus was proved for the first time by in vivo physiological analysis. The identification of the potentially anti-fungal PtoCXE06 gene also laid a theoretical foundation for promoting the genetic improvement of disease-resistance traits in forest trees.