ABSTRACT Unlike traditional two‐component systems, the Rcs system is an atypical multicomponent phosphorelay. It represents one of the most complex signalling networks in bacteria, regulating vital physiological processes; however, its function in the poplar canker bacterium Lonsdalea populi remains unclear. By combining RNA sequencing, electrophoretic mobility shift assays and pathogenicity assays, we identified two novel RcsB target genes: the pectate lyase gene pelE and the TolC‐family outer membrane channel gene omeP . Unphosphorylated RcsB can directly regulate pelE expression to promote host pectin degradation. As PelE degrades the plant cell wall, it activates host immune responses and triggers a burst of reactive oxygen species (ROS). We propose that the transmembrane kinase RcsC senses these stimuli and, via RcsD, transfers phosphate to RcsB, consistent with the conserved Rcs phosphorelay. Phosphorylated RcsB enhanced binding affinity, thereby strengthening transcriptional regulation of omeP and enhancing the tolerance of the pathogen to ROS. This study shows that the Rcs system coordinately regulates omeP and pelE expression to establish an infection pathway, hence promoting L. populi colonization. It highlights a core strategy of woody plant pathogens to degrade host barriers and withstand stress, offering new insights for targeted control.
Rice foot rot, caused by Dickeya oryzae, is a destructive bacterial disease. The Vfm quorum-sensing (QS) system is a key virulence regulator of this pathogen. However, the mechanisms by which it perceives and integrates upstream environmental signals for precise transcriptional control are poorly understood. Here, we identify a cascade regulatory network involving the MarR-family transcription factors OhrR and EmrR. Deletion of ohrR in D. oryzae EC1 significantly reduced the Vfm signalling activity, diminished the production of virulence factors including phytotoxin zeamines and plant cell wall-degrading enzymes, and attenuated its pathogenicity. Molecular analyses revealed that OhrR directly bound to the key promoters in the vfm cluster (PvfmA, PvfmK, PvfmE and PvfmF) and activated their transcription. Intriguingly, OhrR also directly activated the expression of emrR, which encodes another MarR-family transcriptional regulator. In contrast to the activator role of OhrR, EmrR acted as a feedback inhibitor, directly bound to the same vfm promoter regions but repressed their transcription. OhrR-EmrR thus acts as a two-tiered 'activation-inhibition' regulatory circuit in modulation of the Vfm QS system, in which OhrR functions as a master switch for environmental signal response and initiation of the Vfm system, whereas EmrR acts as a precise brake to prevent overexpression of virulence determinants. The findings from this study unveil the fascinating interaction between OhrR-EmrR and the Vfm QS system, and depict a novel mechanism for precise virulence control in D. oryzae via complex, dynamic transcriptional reprogramming.
Botrytis cinerea triggers destructive grey mould with substantial economic losses, yet the functions of numerous pathogenicity genes remain largely unclear. Here, by screening a T-DNA insertion library, we identified BcARS2, a key component of the Microprocessor complex. BcARS2 is constitutively expressed, localizes to the nucleus, and modulates gene expression. The Delta Bcars2 mutants exhibited impaired vegetative growth, enhanced conidiophore sporulation, increased conidiation, reduced sclerotial size, and increased sensitivity to multiple stresses. Pathogenicity assays showed that different inoculation methods led to distinct virulence phenotypes in Delta Bcars2 mutants. Microscopic observations revealed that although the Delta Bcars2 mutant formed normal appressoria and successfully invaded host cells, its invasive hyphae exhibited limited expansion. Furthermore, exogenous cAMP did not restore the impaired development of infection cushions (ICs), resulting in defective infection. Multitranscriptome analyses demonstrated that BcARS2 positively regulates BcPDI1 and BcCGF1, two key genes responsible for reactive oxygen species (ROS) homeostasis and highly expressed during the IC developmental stage. Correspondingly, deletion of BcARS2 upregulated catalase gene expression, elevated catalase activity, reduced H2O2 accumulation, disturbed ROS balance, and ultimately inhibited IC formation. In addition, most toxin biosynthesis-related genes, which are normally upregulated during IC development, were significantly downregulated in the Delta Bcars2 mutant. Collectively, BcARS2 modulates the virulence of B. cinerea by regulating IC development, vegetative hyphal growth, and stress responses.
Plant-pathogenic fungi secrete small proteins, termed effectors, to reprogramme host metabolism and suppress immune responses during infection. Although transcriptional waves of effector expression have been described in several pathosystems, the cis-regulatory elements underlying their temporal coordination remain largely unknown. Here, we investigate the temporal regulation of effector genes in the biotrophic smut fungus Ustilago maydis, a model organism for fungal plant pathogenesis. By integrating transcriptome reanalysis with comparative promoter motif enrichment across biotrophic fungi, we identify distinct promoter motifs associated with defined infection phases. In U. maydis, three candidate cis-regulatory elements correlate with early, proliferative and late infection stages, respectively. Positional enrichment relative to transcription start sites supports their regulatory relevance. Functional dissection of the native pep1, stp1, and cmu1 promoters revealed strongly context-dependent contributions of these elements and synthetic promoter assays showed that the early-phase motif acts as an activating and the proliferative motif as a repressive element. Together, our findings support a combinatorial cis-regulatory code in which activating and repressive promoter elements are integrated in a promoter-specific manner to shape temporal effector expression during U. maydis infection. These elements provide a framework for engineering synthetic fungal promoters with tunable transcriptional outputs.
ABSTRACT Potato virus Y (PVY) is one of the top five economically important plant viruses in the world that affects several plant species including potato, pepper and tomato. We have combined untargeted metabolomics with protein–protein interaction studies to understand the mechanistic details of PVY‐induced metabolic changes that lead to disease development. For this purpose, PVY necrotic strain, PVY NTN , and Nicotiana benthamiana were used as a model system. Gas chromatography–mass spectrometry (GC–MS)‐based metabolomic profiling revealed that PVY infection significantly alters carbohydrate and amino acid metabolism, with elevated levels of glucose‐6‐phosphate, fructose‐6‐phosphate and key amino acids associated with nucleotide biosynthesis. Using yeast two‐hybrid, bimolecular fluorescence complementation and GST pull‐down assays, we identified a direct interaction between the PVY coat protein (CP) and cytosolic phosphoglucomutase (cPGM), a central enzyme regulating cytosolic sugar‐phosphate partitioning among different pathways. Subcellular localization analyses showed that PVY CP localizes to the cytosol, forms aggregates and alters cPGM distribution, recruiting it into CP‐associated cytosolic aggregates. Silencing of cPGM restricted PVY accumulation and disease development, indicating that cPGM is required for efficient PVY infection and contributes to disease development in N. benthamiana . Based on these findings, we propose a working model in which PVY CP hijacks cPGM to redirect carbohydrate flux towards respiratory and pentose phosphate pathways, thereby increasing the availability of energy, reducing equivalents, and biosynthetic precursors required for viral replication. Together, this study provides mechanistic insight into PVY‐induced metabolic reprogramming and identifies cPGM as a key host factor linking viral infection to primary metabolism.
