Characterization of non-canonical G beta-like protein FvGbb2 and its relationship with heterotrimeric G proteins in Fusarium verticillioides

Abstract

Summary Fusarium verticillioides is a fungal pathogen that is responsible for maize ear rot and stalk rot diseases worldwide. The fungus also produces carcinogenic mycotoxins, fumonisins on infested maize. Unfortunately, we still lack clear understanding of how the pathogen responds to host and environmental stimuli to trigger fumonisin biosynthesis. The heterotrimeric G protein complex, consisting of canonical Gα, Gβ and Gγ subunits, is involved in transducing signals from external stimuli to regulate downstream signal transduction pathways. Previously, we demonstrated that Gβ protein FvGbb1 directly impacts fumonisin regulation but not other physiological aspects in F. verticillioides. In this study, we identified and characterized a RACK1 (Receptor for Activated C Kinase 1) homolog FvGbb2 as a putative Gβ‐like protein in F. verticillioides. The mutant exhibited severe defects not only in fumonisin biosynthesis but also vegetative growth and conidiation. FvGbb2 was positively associated with carbon source utilization and stress agents but negatively regulated general amino acid control. While FvGbb2 does not interact with canonical G protein subunits, it may associate with diverse proteins in the cytoplasm to regulate vegetative growth, virulence, fumonisin biosynthesis and stress response in F. verticillioides.

Distinct function of Mediator subunits in fungal development, stress response and secondary metabolism in fungal pathogenFusarium verticillioides

“…To test the role Mediator subunits play in F. verticillioides virulence, we used maize seedling infection assay as previously described (34). After a 10-day incubation at room temperature, all mutants exhibited reduced virulence when compared to the wild-type progenitor.…”

“…F. verticillioides wild-type strain M3125 was used for the construction of the derived mutants used in this study (34). All strains used in this study were grown and evaluated at 28°C on V8, 0.2×PDA and Myro agar plates for mycelial growth (34).…”

“…F. verticillioides wild-type strain M3125 was used for the construction of the derived mutants used in this study (34). All strains used in this study were grown and evaluated at 28°C on V8, 0.2×PDA and Myro agar plates for mycelial growth (34). For conidia gemination assay, conidia (10 6 ) grown on V8 plates for 6 days were harvested and subsequently cultivated in water agar plates for 6.5 h at 28°C in dark.…”

“…All gene deletion mutants were constructed using our standard split-marker gene knock out protocol as previously described (34). Hygromycin B phosphotransferase gene (HPH) was split into two fragments PH (929 bp) and HP (765 bp) and fused with left and right flanking sequence fragments of each targeted gene, respectively (Fig.…”

“…We confirmed targeted gene replacement by PCR, and further tested the absence of gene transcription by qPCR. FvMed1-GFP fusion cassette was constructed and transformed into F. verticillioides following our standard protocol described previously (34). All transformants were verified by PCR and observed under an Olympus BX51 microscope (Olympus America, Melville, NY, USA).…”