Starvation-induced cell fusion and heterokaryosis frequently escape imperfect allorecognition systems to enable parasexual interactions in an asexual fungal pathogen

Abstract

Asexual fungi include important pathogens of plants and other organisms, and their effective management requires understanding of their evolutionary dynamics. Genetic recombination is critical for species adaptability and could be achieved via heterokaryosis and the parasexual cycle in asexual fungi. Here, we investigate the extent and mechanisms of heterokaryosis in the asexual plant pathogen Verticillium dahliae. We used live-cell imaging and genetic complementation assays of tagged V. dahliae strains to analyze the extent of nonself vegetative fusion, heterokaryotic cell fate and nuclear behavior. An efficient CRISPR/Cas9-mediated system was developed to investigate the involvement of autophagy in heterokaryosis. Under starvation, nonself fusion of germinating spores occurs frequently regardless of the previously assessed vegetative compatibility of the partners. Supposedly "incompatible" fusions often establish viable heterokaryotic cells and mosaic mycelia, where nuclei can engage in fusion or transfer of genetic material. The molecular machinery of autophagy has a protective function against destruction of "incompatible" heterokaryons. Our results suggest an autophagy-mediated trade-off between parasexual interactions for genetic exchange and allorecognition systems possibly for mycelial protection from parasitic elements. Our study reveals unexpected capacity for heterokaryosis in V. dahliae and suggests, therefore, important roles of parasexuality in the evolution of asexual fungi.

The NADPH Oxidase A of Verticillium dahliae is Essential for Pathogenicity, Normal Development, and Stress Tolerance, and it Interacts with Yap1 to Regulate Redox Homeostasis

“…Conidial and hyphal fusion is an integral component of the establishment and development of fungal colonies [65], and nonself fusion could possibly grant access to parasexual generation of diversity [23,54]. We have previously demonstrated that NoxA is essential for CAT-mediated fusion of V. dahliae conidia or germlings [23], and we further found in the present study that the requirement for functional NoxA applies to both interacting cells.…”

“…Fusion of vegetatively incompatible strains of fungi, including V. dahliae , usually triggers an incompatibility reaction that can cause cell death of the anastomosed compartments [54]. Based on the observations that this reaction in N. crassa is characterized by induction of ROS [57], and that genes involved in the OSR are up-regulated during CAT formation in Colletotrichum gloeosporioides [58], we used NBT staining to study O 2 •− accumulation in incompatible V. dahliae fusions (pairing Ls.17 H1-sgfp × 123V).…”

“…We used our previously optimized and described methods for reproducible quantification of CAT-mediated self and nonself fusion [23,54] to investigate the possible involvement of noxA and yap1 in somatic cell fusion. Briefly, conidia from 7-day-old PDA cultures were collected and transferred to 6-mm petri dishes, with coverslips at their bottom (containing 5 mL of CAT medium each) to a final concentration of 2.0 × 10 6 conidia/mL.…”