Functional characterization of cytochrome P450 monooxygenases in the cereal head blight fungus Fusarium graminearum

Abstract

Fusarium graminearum is a prominent plant pathogenic fungus causing Fusarium head blight in major cereal crops worldwide. To understand the molecular mechanisms underlying fungal development and virulence, large collections of F. graminearum mutants have been constructed. Cytochrome P450 monooxygenases (P450s) are widely distributed in organisms and are involved in a diverse array of molecular/metabolic processes; however, no systematic functional analysis of P450s has been attempted in filamentous fungi. In this study, we constructed a genome-wide deletion mutant set covering 102 P450s and analyzed these mutants for changes in 38 phenotypic categories, including fungal development, stress responses and responses to several xenobiotics, to build a comprehensive phenotypic dataset. Most P450 mutants showing defective phenotypes were impaired in a single phenotypic trait, demonstrating that our mutant library is a good genetic resource for further fungal genetic studies. In particular, we identified novel P450s specifically involved in virulence (5) and both asexual (1) and sexual development (2). Most P450s seem to play redundant roles in the degradation of xenobiotics in F. graminearum. This study is the first phenome-based functional analysis of P450s, and it provides a valuable genetic resource for further basic and applied biological research in filamentous fungi and other plant pathogens.

Fungal Cytochrome P450s and the P450 Complement (CYPome) of Fusarium graminearum

“…In particular, filamentous fungi have an outstanding ability to degrade a variety of toxic substances (e.g., environmental pollutants, xenobiotics, and plant-derived toxins) [ 16 , 17 , 18 , 19 ], and some filamentous fungi are well known for production of characteristic toxins via CYPs. Recent genetic evidence suggests that CYP enzyme reactions are closely involved in fungal developmental processes and pathogenesis [ 20 , 21 ].…”

“…Four CYPs ( TRI1 , TRI4 , TRI13 , and TRI11 ) are involved in trichothecene biosynthesis [ 71 , 162 ], and Fg08079 and CLM2 are required for the biosynthesis of butenolide [ 163 ] and culmorin [ 164 ], respectively. To systematically characterize the CYPome of F. graminearum , CYP gene knockout mutants were generated, and a comprehensive phenome of 102 CYP mutant library of F. graminearum was tested in 38 traits ( Figure 3 ) [ 20 ]. Notably, specific CYP genes have been identified that are required for virulence (five CYPs), conidiation (one CYP), and sexual development (two CYPs) in this fungus.…”

“…Likewise, some putative xenobiotic-metabolizing CYPs in F . graminearum have been characterized based on genetic evidence [ 20 ]. Fg01972 (CYP505A7) mutant strains showed reduced growth on 1-dodecanol-amended medium compared to the wild type ( Figure 3 a), and transcription of Fg01972 was highly up-regulated in response to exogenous treatment with 1-dodecanol.…”

“…The Fg01972 enzyme may play an important role in detoxification and/or mineralization of this compound. Moreover, most CYP genes (93 of 102) were markedly induced in at least one xenobiotic condition [ 20 ], demonstrating that many are closely related to xenobiotic metabolism, with redundant functions. Further studies are required to identify the substrates of these CYP enzymes.…”

“…Both the CYP505 ( Fg07596 and Fg01972 ) and CYP540 ( Fg02138 and Fg02929 ) families in F . graminearum are highly induced by aliphatics such as n-dodecane and 1-dodecanol [ 20 ]. In addition, Fg10451 belongs to the CYP53A subfamily hydroxylates benzoic acid and other monosubstituted benzoate derivatives, forming para-hydroxylated products in the ascomycete fungi A. niger , A. nidulans, and C. lunatus, as well as in the basidiomycete fungus P. chrysosporium [ 51 , 52 , 55 , 56 ].…”