A transcriptional activator effector of Ustilago maydis regulates hyperplasia in maize during pathogen-induced tumor formation.

Ustilago maydis causes common smut in maize, which is characterized by tumor formation in aerial parts of maize. Tumors result from the de novo cell division of highly developed bundle sheath and subsequent cell enlargement. However, the molecular mechanisms underlying tumorigenesis are still largely unknown. Here, we characterize the U. maydis effector Sts2 (Small tumor on seedlings 2), which promotes the division of hyperplasia tumor cells. Upon infection, Sts2 is translocated into the maize cell nucleus, where it acts as a transcriptional activator, and the transactivation activity is crucial for its virulence function. Sts2 interacts with ZmNECAP1, a yet undescribed plant transcriptional activator, and it activates the expression of several leaf developmental regulators to potentiate tumor formation. On the contrary, fusion of a suppressive SRDX-motif to Sts2 causes dominant negative inhibition of tumor formation, underpinning the central role of Sts2 for tumorigenesis. Our results not only disclose the virulence mechanism of a tumorigenic effector, but also reveal the essential role of leaf developmental regulators in pathogen-induced tumor formation.

Maize Phytocytokines Modulate Pro-Survival Host Responses and Pathogen Resistance.

Phytocytokines are signaling peptides that alert plant cells of danger. However, the downstream responses triggered by phytocytokines and their effect on plant survival are still largely unknown. Here, we have identified three biologically active maize orthologues of phytocytokines previously described in other plants. The maize phytocytokines show common features with microbe-associated molecular patterns (MAMPs), including the induction of immune-related genes and activation of papain-like cysteine proteases. In contrast to MAMPs, phytocytokines do not promote cell death in the presence of wounding. In infection assays with two fungal pathogens, we found that phytocytokines affect the development of disease symptoms, likely due to the activation of phytohormonal pathways. Collectively, our results show that phytocytokines and MAMPs trigger unique and antagonistic features of immunity. We propose a model in which phytocytokines activate immune responses partially similar to MAMPs but, in contrast to microbial signals, they act as danger and survival molecules to the surrounding cells. Future studies will focus on the components determining the divergence of signaling outputs upon phytocytokine activation. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Host apoplastic cysteine protease activity is suppressed during the mutualistic association of Lolium perenne and Epichloë festucae.

Plants secrete various defence-related proteins into the apoplast, including proteases. Papain-like cysteine proteases (PLCPs) are central components of the plant immune system. To overcome plant immunity and successfully colonize their hosts, several plant pathogens secrete effector proteins inhibiting plant PLCPs. We hypothesized that not only pathogens, but also mutualistic microorganisms interfere with PLCP-meditated plant defences to maintain endophytic colonization with their hosts. Epichloë festucae forms mutualistic associations with cool season grasses and produces a range of secondary metabolites that protect the host against herbivores. In this study, we performed a genome-wide identification of Lolium perenne PLCPs, analysed their evolutionary relationship, and classified them into nine PLCP subfamilies. Using activity-based protein profiling, we identified four active PLCPs in the apoplast of L. perenne leaves that are inhibited during endophyte interactions. We characterized the L. perenne cystatin LpCys1 for its inhibitory capacity against ryegrass PLCPs. LpCys1 abundance is not altered during the mutualistic interaction and it mainly inhibits LpCP2. However, since the activity of other L. perenne PLCPs is not sensitive to LpCys1, we propose that additional inhibitors, likely of fungal origin, are involved in the suppression of apoplastic PLCPs during E. festucae infection.

Comparative transcriptome profiling identifies maize line specificity of fungal effectors in the maize-Ustilago maydis interaction.

The biotrophic pathogen Ustilago maydis causes smut disease on maize (Zea mays) and induces the formation of tumours on all aerial parts of the plant. Unlike in other biotrophic interactions, no gene-for-gene interactions have been identified in the maize-U. maydis pathosystem. Thus, maize resistance to U. maydis is considered a polygenic, quantitative trait. Here, we study the molecular mechanisms of quantitative disease resistance (QDR) in maize, and how U. maydis interferes with its components. Based on quantitative scoring of disease symptoms in 26 maize lines, we performed an RNA sequencing (RNA-Seq) analysis of six U. maydis-infected maize lines of highly distinct resistance levels. The different maize lines showed specific responses of diverse cellular processes to U. maydis infection. For U. maydis, our analysis identified 406 genes being differentially expressed between maize lines, of which 102 encode predicted effector proteins. Based on this analysis, we generated U. maydis CRISPR/Cas9 knock-out mutants for selected candidate effector sets. After infections of different maize lines with the fungal mutants, RNA-Seq analysis identified effectors with quantitative, maize line-specific virulence functions, and revealed auxin-related processes as a possible target for one of them. Thus, we show that both transcriptional activity and virulence function of fungal effector genes are modified according to the infected maize line, providing insights into the molecular mechanisms underlying QDR in the maize-U. maydis interaction.