Effect of ChCUC1 induction on gene expression in Cardamine hirsuta leaf development

Here we investigate the function of CUC1(CUP-SHAPED COTYLEDON1) in the diversification of leaf forms between simple-leaved Arabidopsis thaliana and compound-leaved Cardamine hirsuta. CUC transcription factors are conserved regulators in leaf margin dissection and leaflet formation. ChCUC1, ChCUC2 and ChCUC3 function redundantly and are required for the leaflet formation in C. hirsuta. Recently we discovered that ChCUC1 has species species-specific expression in young leaves of C.hirsuta. Moreover, interspecies gene transfer of ChCUC1 allele into A.thaliana is sufficient to increase leaf complexity. On this basis, we hypothesize that redeployment of ChCUC1 in leaves contributes to the formation of leaflets instead of serrations. However, the mechanism underlying ChCUC1 regulating cell division, cell polarity, cytoskeleton and thus leaf marginal patterning remains elusive. To this end, we make use of chromatin immunoprecipitation sequencing(ChIP-seq), transcriptomic, comparative genetics and advanced imaging approaches to identify the downstream regulating genes of ChCUC1. To this end, we generated RCOp::LhG4:GR; Op::ChCUC1:Venus transgenic line in C. hirsuta. We harvested 12-days-old seedlings harboring RCOp::LhG4:GR; Op::ChCUC1:Venus, which were incubated in liquid MS medium with 10 μM Dexamethasone (induction) or 0.1% DMSO (control). Seedlings without cotyledons, roots, and mature leaves were harvested at 2, 4, 6, and 8 hours after treatment. For each replicate, twenty seedlings were collected for RNA isolation. Three independent replicates treated with Dexamethasone were used as the experimental group, and three independent replicates treated with DMSO were used as the control group.