Bioinformatics Prediction of Polyketide Synthase Gene Clusters from Mycosphaerella fijiensis

Abstract

Mycosphaerella fijiensis, causal agent of black Sigatoka disease of banana, is a Dothideomycete fungus closely related to fungi that produce polyketides important for plant pathogenicity. We utilized the M. fijiensis genome sequence to predict PKS genes and their gene clusters and make bioinformatics predictions about the types of compounds produced by these clusters. Eight PKS gene clusters were identified in the M. fijiensis genome, placing M. fijiensis into the 23rd percentile for the number of PKS genes compared to other Dothideomycetes. Analysis of the PKS domains identified three of the PKS enzymes as non-reducing and two as highly reducing. Gene clusters contained types of genes frequently found in PKS clusters including genes encoding transporters, oxidoreductases, methyltransferases, and non-ribosomal peptide synthases. Phylogenetic analysis identified a putative PKS cluster encoding melanin biosynthesis. None of the other clusters were closely aligned with genes encoding known polyketides, however three of the PKS genes fell into clades with clusters encoding alternapyrone, fumonisin, and solanapyrone produced by Alternaria and Fusarium species. A search for homologs among available genomic sequences from 103 Dothideomycetes identified close homologs (>80% similarity) for six of the PKS sequences. One of the PKS sequences was not similar (< 60% similarity) to sequences in any of the 103 genomes, suggesting that it encodes a unique compound. Comparison of the M. fijiensis PKS sequences with those of two other banana pathogens, M. musicola and M. eumusae, showed that these two species have close homologs to five of the M. fijiensis PKS sequences, but three others were not found in either species. RT-PCR and RNA-Seq analysis showed that the melanin PKS cluster was down-regulated in infected banana as compared to growth in culture. Three other clusters, however were strongly upregulated during disease development in banana, suggesting that they may encode polyketides important in pathogenicity.

Transcriptome sequencing of Mycosphaerella fijiensis during association with Musa acuminata reveals candidate pathogenicity genes

“…2 and 7) [36]. In addition to the three polyketide synthase gene clusters recently described [17], we showed that an NRPS, an NRPS-like, and a fusicoccane gene cluster had higher expression in infected leaf tissue compared to medium (Fig. 7).…”

Progress in understanding Pseudocercospora banana pathogens and the development of resistant Musa germplasm

“…Furthermore, a bioinfomatic approach to predict polyketide synthase ( PKS ) gene clusters from P. fijiensis found that the fungus contained three PKS genes that were also present in P. musae and P. eumusae but absent in other fungal species. Further comparison to P. musae and P. eumusae PKS gene clusters showed that many of the PKS genes were similar to those of P. fijiensis , with a single gene exception that encoded a unique compound (Noar & Daub, ). An analysis of several isolates of P. fijiensis has also revealed allelic variation in some of the effector‐coding sequences (Stergiopoulos et al ., ).…”

“…Such findings, coupled with the current understanding that the three pathogens have different strategies for manipulating the host immune system, have great implications especially for the development of Musa germplasm with resistance to Pseudocercospora banana pathogens. Recently, transcriptomic and bioinformatic analyses have identified genes that code for molecules that aid Pseudocercospora banana pathogens to successfully infect banana, including PKS and proteins involved in metabolism and enzymatic degradation (Chang et al ., ; Noar & Daub, ,b). Bioinformatic analysis has also demonstrated that some of the effectors are unique to the three Pseudocercospora banana pathogens while one is only present in P. fijiensis and not in any known fungal pathogen (Noar & Daub, ).…”

“…It is suspected that microRNAs, metabolites and toxins play effector roles in virulence of Pseudocercospora banana pathogens, although this has not been clearly demonstrated (Weiberg et al ., ). Few effector proteins that are functional orthologues of the much studied Cladosporium fulvum have been identified in the genome of P. musae and P. fijiensis (Stergiopoulos et al ., ; Passos et al ., ; Arango‐Isaza et al ., ; Noar & Daub, ). Notably, a higher number of effectors have been identified in P. fijiensis than other related fungi such as M. graminicola , P. eumusae , P. musae and C. fulvum (Cho et al ., ; Arango‐Isaza et al ., ; Friesen, ; Noar & Daub, ).…”

“…Indeed, bioinformatics analysis of the repetitive sequences has revealed that gene expansions and increases in repetitive sequences lead to emergence of new strains/pathotypes of P. fijiensis (Santana et al ., ; Möller & Stukenbrock, ). To some extent these genomic expansions could be responsible for several PKS genes that are only found in the Pseudocercospora banana pathogens and no other fungal genera, making it an even more complex pathosystem (Noar & Daub, ).…”