Testing the impact of DNA methylation in periderm development

The periderm has a protective role against adverse environmental conditions and is mainly made up of suberized phellem or cork cells produced by the phellogen. The cork tissue is interrupted by lenticels (cork porosity) produced by the lenticular phellogen, that allow gas exchange between the external environment and the inner tissues. When present at high levels, the lenticels strongly depreciate cork industrial performance and economical value. Studies in cork oak, a species that produces an exceptionally thick periderm due to its highly active and long-living phellogen, evidenced an increase in de novo DNA methylation during phellem differentiation and DNA methylation polymorphisms statistically associated with lenticel characters, suggesting a potential role of DNA methylation in the control of phellogen-daughter cells specification. To test this hypothesis and characterize the impacts of DNA methylation in periderm development, we are using potato tuber periderm as a model and an inducible genome editing (IGE) system, in which a highly active promoter in the lenticular phellogen drives CRISPR/Cas9, to generate loss-of-function mutants of DNA methyltransferase genes. IGE constructs targeting StMET1, StDRM1, StDRM2, StCMT2 and StCMT3 genes, and two double mutants targeting StDRM1DRM2 and StCMT2CMT3 have been successfully generated. The constructs targeting StDRM1 and StCMT2 were already transferred to potato plants to be further induced in microtubers’ periderm. Histological characterization of the mutant’s periderm complemented with transcriptomic and methylome analyses will clarify the role of DNA methylation in cork formation and provide insights of the underlying epigenetic mechanisms.