Plant leaf margins produce small outgrowths or teeth causing serration in a regular arrangement, which is specified by auxin maxima. In Arabidopsis, the spatiotemporal pattern of auxin dependents on both, the transcription factor CUC2 and the signal peptide EPFL2, a ligand of the growth-promoting receptor kinase ERECTA (ER). Ectopic expression of CUC2 can have contrary effects on leaf growth. Ubiquitous expressed CUC2 suppresses growth in the whole leaf, whereas cuc2-1D mutants have enlarged leaves, through ER-dependent cell proliferation in the teeth. Here we investigated the growth dynamics of cuc2-1D leaves and the growth restricting the function of CUC2 using the ubiquitous inducible CUC2-GR transgene. In time courses, we dissected the serration promoting the function of CUC2 in the leaf margin and ectopic growth inhibition by CUC2 in the leaf plate. We found that CUC2 limits growth rather by cell cycle inhibition than by cell size control. Furthermore, endogenous CUC2 was rapidly induced by CUC2-GR indicating a possible auto-inducible feedback. In contrast, EPFL2 was quickly decreased by transient CUC2 induction but increased in cuc2-3 mutant leaves suggesting that CUC2 can also counteract the EPFL2-ER pathway. Therefore, tooth growth promotion and growth inhibition by CUC2 involve partially the same mechanism but in contrary ways.

Polycomb group (PcG) protein-mediated histone methylation (H3K27me3) controls the correct spatiotemporal expression of numerous developmental regulators in Arabidopsis. Epigenetic silencing of the stem cell factor gene WUSCHEL (WUS) in floral meristems (FMs) depends on H3K27me3 deposition by PcG proteins. However, the role of H3K27me3 in silencing of other meristematic regulator and pluripotency genes during FM determinacy has not yet been studied. To this end, we report the genome-wide dynamics of H3K27me3 levels during FM arrest and the consequences of strongly depleted PcG activity on early flower morphogenesis including enlarged and indeterminate FMs. Strong depletion of H3K27me3 levels results in misexpression of the FM identity gene AGL24, which partially causes floral reversion leading to ap1-like flowers and indeterminate FMs ectopically expressing WUS and SHOOT MERISTEMLESS (STM). Loss of STM can rescue supernumerary floral organs and FM indeterminacy in H3K27me3-deficient flowers, indicating that the hyperactivity of the FMs is at least partially a result of ectopic STM expression. Nonetheless, WUS remained essential for the FM activity. Our results demonstrate that PcG proteins promote FM determinacy at multiple levels of the floral gene regulatory network, silencing initially floral regulators such as AGL24 that promotes FM indeterminacy and, subsequently, meristematic pluripotency genes such as WUS and STM during FM arrest.

Plants have the potency to regenerate adventitious roots from aerial organs after detachment. In Arabidopsis thaliana, de novo root regeneration (DNRR) from leaf explants is triggered by wounding signaling that rapidly induces the expression of the ETHYLENE RESPONSE FACTOR (ERF) transcription factors ERF109 and ABR1 (ERF111). In turn, the ERFs promote the expression of ASA1, an essential enzyme of auxin biosynthesis, which contributes to rooting by providing high levels of auxin near the wounding side of the leaf. Here, we show that the loss of the epigenetic regulator ULTRAPETALA1 (ULT1), which interacts with Polycomb and Trithorax Group proteins, accelerates and reinforces adventitious root formation. Expression of ERF109 and ASA1 was increased in ult1 mutants, whereas ABR1 was not significantly changed. Cultivation of explants on media with exogenous auxin equates adventitious root formation in wild-type with ult1 mutants, suggesting that ULT1 negatively regulates DNRR by suppressing auxin biosynthesis. Based on these findings, we propose that ULT1 is involved in a novel mechanism that prevents overproliferation of adventitious roots during DNRR.

Orchardgrass (Dactylis glomerata) is one of the top four perennial forages worldwide and, despite its large economic advantages, often threatened by various environmental stresses. WRKY transcription factors (TFs) can regulate a variety of plant processes, widely participate in plant responses to biotic and abiotic stresses, and are one of the largest gene families in plants. WRKYs can usually bind W-box elements specifically. In this study, we identified a total of 93 DgWRKY genes and 281 RGAs, including 65, 169 and 47 nucleotide-binding site-leucine-rich repeats (NBS-LRRs), leucine-rich repeats receptor-like protein kinases (LRR-RLKs), and leucine-rich repeats receptor-like proteins (LRR-RLPs), respectively. Through analyzing the expression of DgWRKY genes in orchardgrass under different environmental stresses, it was found that many DgWRKY genes were differentially expressed under heat, drought, submergence, and rust stress. In particular, it was found that the greatest number of genes were differentially expressed under rust infection. Consistently, GO and KEGG enrichment analysis of all genes showed that 78 DgWRKY TFs were identified in the plant–pathogen interaction pathway, with 59 of them differentially expressed. Through cis-acting element prediction, 154 RGAs were found to contain W-box elements. Among them, DG6C02319.1 (a member of the LRR-RLK family) was identified as likely to interact with 14 DGWRKYs. Moreover, their expression levels in susceptible plants after rust inoculation were first up-regulated and then down-regulated, while those in the resistant plants were always up-regulated. In general, DgWRKYs responded to both biotic stress and abiotic stress. DgWRKYs and RGAs may synergistically respond to the response of orchardgrass to rust. This study provides meaningful insight into the molecular mechanisms of WRKY proteins in orchardgrass.

Adventitious root (AR) formation in plants originates from non-root organs such as leaves and hypocotyls. Auxin signaling is essential for AR formation, but the roles of other phytohormones are less clear. In Arabidopsis, at least two distinct mechanisms can produce ARs, either from hypocotyls as part of the general root architecture or from wounded organs during de novo root regeneration (DNRR). In previous reports, gibberellin acid (GA) appeared to play reverse roles in both types of ARs, since GA treatment blocks etiolation-induced AR formation from hypocotyls, whereas GA synthesis and signaling mutants apparently displayed reduced DNRR from detached leaves. In order to clarify this contradiction, we employed the GA biosynthesis inhibitor paclobutrazol (PBZ) and found that PBZ had positive effects on both types of AR formation in Arabidopsis. Consistently, GA treatment had negative effects on both AR formation mechanisms, while loss of GA synthesis and signaling promoted DNRR under our conditions. Our results show that PBZ treatment can rescue declined AR formation in difficult-to-root leaf explants such as erecta receptor mutants. Furthermore, transcriptional profiling revealed that PBZ treatment altered GA, brassinosteroids, and auxin responses, which included the up-regulation of LBD16 that is well known for its pivotal role in AR initiation.