The RasGEF FgCdc25 regulates fungal development and virulence in Fusarium graminearum via cAMP and MAPK signalling pathways.

Ras GTPases act as molecular switches to control various cellular processes by coupling integrated signals in eukaryotes. Activities of Ras GTPases are triggered by Ras GTPase guanine nucleotide exchange factors (RasGEFs) in general, whereas the role of RasGEF in plant pathogenic fungi is largely unknown. In this study, we characterized the only RasGEF protein in Fusarium graminearum, FgCdc25, by combining genetic, cytological and phenotypic strategies. FgCdc25 directly interacted with RasGTPase FgRas2, but not FgRas1, to regulate growth and sexual reproduction. Mutation of the FgCDC25 gene resulted in decreased toxisome formation and deoxynivalenol (DON) production, which was largely depended on cAMP signalling. In addition, FgCdc25 indirectly interacted with FgSte11 in FgSte11-Ste7-Gpmk1 cascade, and the ΔFgcdc25 strain totally abolished the formation of infection structures and was nonpathogenic in planta, which was partially recovered by addition of exogenous cAMP. In contrast, FgCdc25 directly interplayed with FgBck1 in FgBck1-MKK1-Mgv1 cascade to negatively control cell wall integrity. Collectively, these results suggest that FgCdc25 modulates cAMP and MAPK signalling pathways and further regulates fungal development, DON production and plant infection in F. graminearum.

Contribution of peroxisomal docking machinery to mycotoxin biosynthesis, pathogenicity and pexophagy in the plant pathogenic fungus Fusarium graminearum.

Peroxisomal proliferation is highly stimulated during the biosynthesis of mycotoxins and plant infection by Fusarium graminearum. Currently, the functions of the peroxisome in these cellular processes are poorly understood. In this study, we applied genetic, cell biological and biochemical analyses to investigate the functions of the peroxisomes. We constructed targeted deletion of docking machinery components, including FgPex13, FgPex14 and the filamentous fungal specific peroxin FgPex33. Our results indicated that peroxisome dysfunction resulted in a shortage of acetyl-CoA, the precursor of trichothecene biosynthesis, and subsequently decreased deoxynivalenol (DON) production. Deletion mutants of ΔFgPex13, ΔFgPex14 or ΔFgPex33 showed an increased accumulation of endogenous reactive oxygen species (ROS) and reduced phosphorylation of MAP (Mitogen-Activated Protein) kinase FgMgv1. In addition, mutants of the docking peroxin exhibited increased sensitivity toward host oxidative bursts and cell wall integrity stress agents and reduced virulence on host plants. More importantly, we found for the first time that FgPex14 is required for pexophagy in F. graminearum. Overall, our study suggests that peroxisomes play critical roles in DON biosynthesis and virulence in F. graminearum.

Efficacy of Nifekalant in Patients With Wolff-Parkinson-White Syndrome and Atrial Fibrillation: Electrophysiological and Clinical Findings.

Background The efficacy of nifekalant in preexcited atrial fibrillation ( AF ) has not been assessed. Methods and Results The study populations consisted of patients with sustained preexcited AF (n=51), paroxysmal supraventricular tachycardia (n=201), and persistent AF (n=87). Effects of intravenous infusion of nifekalant were assessed on electrophysiological and clinical parameters. Nifekalant prolonged the shortest preexcited R-R, the average preexcited R-R, and the average R-R intervals from 290±35 to 333±44 ms, 353±49 to 443±64 ms, and 356±53 to 467±75 ms, respectively, in patients with preexcited AF (all P<0.001). Nifekalant also decreased the percentage of preexcited QRS complexes, heart rate, and increased systolic pressure (all P<0.001). Nifekalant terminated AF in 33 of 51 patients (65%). Similar effects were also observed in a subgroup of 12 patients with preexcited AF and impaired left ventricular function. In patients with paroxysmal supraventricular tachycardia, nifekalant significantly prolonged the effective refractory period, the block cycle length of the antegrade accessory pathway, and the atrial effective refractory period (all P<0.001). Nifekalant had no effect on the effective refractory period of the antegrade atrioventricular node. Finally, in patients with persistent AF without an accessory pathway, nifekalant did not significantly decrease the ventricular rate of AF . One patient developed Torsades de Pointes. No other adverse effects were observed. Conclusions Nifekalant prolongs the effective refractory period of the antegrade accessory pathway and atrium without blocking antegrade conduction through the atrioventricular node, leading to slowing and/or to termination of preexcited AF . Thus, nifekalant might be an effective and a relatively safe drug in patients with preexcited AF .

The mitochondrial membrane protein FgLetm1 regulates mitochondrial integrity, production of endogenous reactive oxygen species and mycotoxin biosynthesis in Fusarium graminearum.

Deoxynivalenol (DON) is a mycotoxin produced in cereal crops infected with Fusarium graminearum. DON poses a serious threat to human and animal health, and is a critical virulence factor. Various environmental factors, including reactive oxygen species (ROS), have been shown to interfere with DON biosynthesis in this pathogen. The regulatory mechanisms of how ROS trigger DON production have been investigated extensively in F. graminearum. However, the role of the endogenous ROS-generating system in DON biosynthesis is largely unknown. In this study, we genetically analysed the function of leucine zipper-EF-hand-containing transmembrane 1 (LETM1) superfamily proteins and evaluated the role of the mitochondrial-produced ROS in DON biosynthesis. Our results show that there are two Letm1 orthologues, FgLetm1 and FgLetm2, in F. graminearum. FgLetm1 is localized to the mitochondria and is essential for mitochondrial integrity, whereas FgLetm2 plays a minor role in the maintenance of mitochondrial integrity. The ΔFgLetm1 mutant demonstrated a vegetative growth defect, abnormal conidia and increased sensitivity to various stress agents. More importantly, the ΔFgLetm1 mutant showed significantly reduced levels of endogenous ROS, decreased DON biosynthesis and attenuated virulence in planta. To our knowledge, this is the first report showing that mitochondrial integrity and endogenous ROS production by mitochondria are important for DON production and virulence in Fusarium species.