The Arf‐GAP Proteins AoAge1 and AoAge2 are Required for Mycelial Growth, Conidiation, Stress Responses, Trap Formation, and the Secondary Metabolism in the Nematode-Trapping Fungus Arthrobotrys oligospora

ADP-ribosylation factors (Arfs) belong to the small GTPases superfamily and regulate mycelial development, endocytosis, and virulence in fungi. Nematode-trapping (NT) fungi can form unique infection structures (traps) to capture and kill free-living nematodes, thus play a potential role in the biocontrol of nematodes. Arthrobotrys oligospora is a representative species of the NT fungi. Here, the functions of two Arf-GAPs, Age1 (AoAge1) and Age2 (AoAge2) orthologue to S. cerevisiae were identified in A. oligospora by gene knockout and multi-phenotypic analysis. It was found that AoAge1 and AoAge2, especially AoAge2, play an important role in vegetative growth, conidiation, trap formation, DNA damage, endoplasmic reticulum stress, mitochondrial activity, endocytosis, heat shock stress, ROS level, and autophagy of A. oligospora. Importantly, our transcriptomic analysis (12 h) showed that nearly 62.7% of A. oligospora genes (11479) were directly or indirectly regulated by AoAge2, indicating that Arf-GAP plays a global role in the growth and development of A. oli gospora. Revealed by transcriptomic analysis differentially upregulated genes in the absence of Aoage2 were involved in autophagy and carbohydrate, amino acid and sphingolipid metabolism while downregulated genes were involved in longevity regulating pathway and carbohydrate, amino acid and secondary metabolites biosynthesis. In addition, metabolomic analysis showed that many compounds were markedly reduction in ΔAoage2 mutants strain. Our results highlighted that AoAge1 and AoAge2 play a crucial role in the asexual growth, development, differentiation, and lifestyle transition, especially, AoAge2 is indispensable for conidiation and trap morphogenesis of A. oligospora. These results demonstrated crucial roles for AoAge1 and AoAge2 in the fungal growth, environmental adaption and pathogenicity, which provided insights into the function of Arf-GAPs in A. oligospora and other fungi. Mycelial samples of the WT and ΔAoage2 mutant strains were collected after trap formation induced by nematodes at different time points (0, 6, 12, and 24 h), and cDNA libraries of each sample were constructed and sequenced.