Ke-Qin Zhang

The human transmembrane protease, serine 2 (TMPRSS2), essential for SARS-CoV-2 entry, is a key antiviral target. Here, we computationally profiled the TMPRSS2-binding affinities of 15 antiviral compounds. Molecular dynamics (MD) simulations for the docked complexes revealed that three compounds exited the substrate-binding cavity (SBC), suggesting noncompetitive inhibition. Of the remaining compounds, five charged ones exhibited reduced binding stability due to competing electrostatic interactions and increased solvent exposure, while seven neutral compounds showed stronger binding affinity driven by van der Waals (vdW) interactions compensating for unfavorable electrostatic effects (including electrostatic interactions and desolvation penalties). Positive and negative hotspot residues were identified as uncharged and charged, respectively, both lining the SBC. Despite forming diverse interactions with compounds, the burial of positive hotspots led to strong vdW interactions that overcompensated for unfavorable electrostatic effects, whereas negative hotspots incurred high desolvation penalties, negating any favorable contributions. Charged residues at the SBC’s outer rim can reduce binding affinity significantly when forming hydrogen bonds or salt bridges. These findings underscore the importance of enhancing vdW interactions with uncharged residues and minimizing the unfavorable electrostatic effects of charged residues, providing valuable insights for designing effective TMPRSS2 inhibitors.

The methylation of lysine 4 of histone H3 (H3K4), catalyzed by the histone methyltransferase KMT2/SET1, has been functionally identified in many pathogenic fungi but remains unexplored in nematode-trapping fungi (NTFs). Here, we report a regulatory mechanism of an H3K4-specific SET1 orthologue, AoSET1, in the typical nematode-trapping fungus Arthrobotrys oligospora. When the fungus is induced by the nematode, the expression of AoSET1 is up-regulated. Disruption of AoSet1 led to the abolishment of H3K4me. Consequently, the yield of traps and conidia of ΔAoSet1 was significantly lower than that of the WT strain, and the growth rate and pathogenicity were also compromised. Moreover, H3K4 trimethylation was enriched mainly in the promoter of two bZip transcription factor genes (AobZip129 and AobZip350) and ultimately up-regulated the expression level of these two transcription factor genes. In the ΔAoSet1 and AoH3^K4A strains, the H3K4me modification level was significantly decreased at the promoter of transcription factor genes AobZip129 and AobZip350. These results suggest that AoSET1-mediated H3KEme serves as an epigenetic marker of the promoter region of the targeted transcription factor genes. Furthermore, we found that AobZip129 negatively regulates the formation of adhesive networks and the pathogenicity of downstream AoPABP1 and AoCPR1. Our findings confirm that the epigenetic regulatory mechanism plays a pivotal role in regulating trap formation and pathogenesis in NTFs, and provide novel insights into the mechanisms of interaction between NTFs and nematodes.

Dear Editor,

Plant-parasitic nematodes cause severe economic losses to agriculture. As important biocontrol agents, nematophagous fungi evolved the ability to obtain nitrogen sources from nematodes. However, the impact of nitrogen sources on the growth and development of these fungi is largely unknown. In this study, we aimed to better understand how nitrogen sources could influence vegetative growth and conidiation through epigenetic regulation in the nematophagous fungus, Purpureocillium lavendulum. Through nutrition screening, we found a phenomenon of the fungus, limited colony extension with a large amount of conidia production when cultured on PDA media, can be altered by adding ammonia nitrate. Characterized by site-directed mutagenesis, the histone H3K14 acetylation was found to be involved in the alternation. Furthermore, the acetyltransferase PlGCN5 was responsible for H3K14 acetylation. Knockout of Plgcn5 severely diminished conidiation in P. lavendulum. Chip-seq showed that H3K14ac distributed in conidiation regulating genes, and genes in the MAPK pathway which may be the downstream targets in the regulation. These findings suggest that histone modification and nitrogen sources coordinated lifestyle regulation in P. lavendulum, providing new insight into the mechanism of growth regulation by nutritional signals for the carnivorous fungus.

Predatory fungi possess intricate signal transduction systems that regulate their development and support successful infection of the host. Herein, we characterized three components of the cell wall integrity-controlling pathway, namely protein kinase C (AoPKC), SLT2-MAPK (AoSLT2), and SWI6 (AoSWI6), in a representative nematode-trapping fungus Arthrobotrys oligospora, using gene disruption and multi-omics approaches. The phenotypic traits (such as mycelia development, conidiation, stress response, and trap morphogenesis) and metabolic profiles of ΔAopkc and ΔAoswi6 mutants were similar but differed from those of the ΔAoslt2 mutants. Transcriptomic analysis indicated that the genes differentially expressed in the absence of Aoswi6 were involved in DNA replication, repair, and recombination during trap formation. Moreover, the yeast two-hybrid assay showed that AoPKC interacted with AoSWI6, suggesting that in A. oligospora, PKC can directly regulate SWI6, bypassing the SLT2 signaling cascade. Conclusively, our findings deepen our understanding of the regulatory mechanism of asexual development and lifestyle switching in nematode-trapping fungi.

Purpureocillium lavendulum is a fungus with promising biocontrol applications. Here, transcriptome data acquired during the infection of Caenorhabditis elegans by Purpureocillium lavendulum showed that the transcription of metabolite synthesis genes was significantly up-regulated after 24 and 48 h of the fungus-nematode interaction. Then, the up-regulated transcription level of lipoxygenase was confirmed by RT-qPCR. The ultra-performance liquid chromatography-mass spectrometry (UPLC-MS) analysis of differential metabolites revealed that this interaction resulted in the emergence of new metabolites or enhanced the production of metabolites. The results of the UPLC-MS analysis and the nematicidal assay were used to establish optimal culturing conditions under which 12 metabolites, including 3 hydroxylated C₁₈ fatty acids and 9 steroids, were isolated and identified. Among them, hydroxylated fatty acids showed pronounced nematicidal activity against Meloidogyne incognita, and two degradative sterols showed chemotaxis activity to M. incognita. This study lays a foundation for the function of lipoxygenase and its products during the infection of Purpureocillium lavendulum.

Small GTPases from the ADP-ribosylation factor (Arf) family and their activating proteins (Arf-GAPs) regulate mycelial development, endocytosis, and virulence in fungi. Here, we identified two orthologous Arf-GAP proteins, AoGcs1 and AoGts1, in a typical nematode-trapping fungus Arthrobotrys oligospora. The transcription of Aogcs1 and Aogts1 was highly expressed in the sporulation stage. The deletion of Aogcs1 and Aogts1 caused defects in DNA damage, endocytosis, scavenging of reactive oxygen species, lipid droplet storage, mitochondrial activity, autophagy, serine protease activity, and the response to endoplasmic reticulum stress. The combined effects resulted in slow growth, decreased sporulation capacity, increased susceptibility to chemical stressors and heat shock, and decreased pathogenicity of the mutants compared with the wild-type (WT) strain. Although deletion of Aogcs1 and Aogts1 produced similar phenotfypic traits, their roles varied in conidiation and proteolytic activity. The ΔAogts1 mutant showed a remarkable reduction in conidial yield compared with the WT strain but not in proteolytic activity; in contrast, the ΔAogcs1 mutant showed an increase in proteolytic activity but not in sporulation. In addition, the growth of ΔAogcs1 and ΔAogts1 mutants was promoted by rapamycin, and the ΔAogts1 mutant was sensitive to H-89. Collectively, the ΔAogts1 mutant showed a more remarkable difference compared with the WT strain than the ΔAogcs1 mutant. Our study further illustrates the importance of Arf-GAPs in the growth, development, and pathogenicity of nematode-trapping fungi.

Nematode-trapping (NT) fungi play a significant role in the biological control of plant- parasitic nematodes. NT fungi, as a predator, can differentiate into specialized structures called “traps” to capture, kill, and consume nematodes at a nutrient-deprived condition. Therefore, trap formation is also an important indicator that NT fungi transition from a saprophytic to a predacious lifestyle. With the development of gene knockout and multiple omics such as genomics, transcriptomics, and metabolomics, increasing studies have tried to investigate the regulation mechanism of trap formation in NT fungi. This review summarizes the potential regulatory mechanism of trap formation in NT fungi based on the latest findings in this field. Signaling pathways have been confirmed to play an especially vital role in trap formation based on phenotypes of various mutants and multi-omics analysis, and the involvement of small molecule compounds, woronin body, peroxisome, autophagy, and pH-sensing receptors in the formation of traps are also discussed. In addition, we also highlight the research focus for elucidating the mechanism underlying trap formation of NT fungi in the future.

Phosphodiesterases are essential regulators of cyclic nucleotide signaling with diverse physiological functions. Two phosphodiesterases, PdeH and PdeL, have been identified from yeast and filamentous fungi. Here, the orthologs of PdeH and PdeL were characterized in a typical nematode-trapping fungus Arthrobotrys oligospora by gene disruption and phenotypic comparison. Deletion of AopdeH caused serious defects in mycelial growth, conidiation, stress response, trap formation, and nematicidal efficiency compared to the wild-type strain. In contrast, these phenotypes have no significant difference in the absence of AopdeL. In addition, deletion of AopdeH and AopdeL resulted in a remarkable increase in cAMP level during vegetative growth and trap formation, and the number of autophagosomes was decreased in ΔAopdeH and ΔAopdeL mutants, whereas their volumes considerably increased. Moreover, metabolomic analyses revealed that many metabolites were downregulated in ΔAopdeH mutant compared to their expression in the wild-type strain. Our results indicate that AoPdeH plays a crucial role in mycelial growth, conidiation, stress response, secondary metabolism, and trap formation. In contrast, AoPdeL only plays a minor role in hyphal and conidial morphology, autophagy, and trap formation in A. oligospora. This work expands the roles of phosphodiesterases and deepens the understanding of the regulation of trap formation in nematode-trapping fungi.