A Novel Type Pathway-Specific Regulator and Dynamic Genome Environments of a Solanapyrone Biosynthesis Gene Cluster in the Fungus Ascochyta rabiei

Abstract

c Secondary metabolite genes are often clustered together and situated in particular genomic regions, like the subtelomere, that can facilitate niche adaptation in fungi. Solanapyrones are toxic secondary metabolites produced by fungi occupying different ecological niches. Full-genome sequencing of the ascomycete Ascochyta rabiei revealed a solanapyrone biosynthesis gene cluster embedded in an AT-rich region proximal to a telomere end and surrounded by Tc1/Mariner-type transposable elements. The highly AT-rich environment of the solanapyrone cluster is likely the product of repeat-induced point mutations. Several secondary metabolism-related genes were found in the flanking regions of the solanapyrone cluster. Although the solanapyrone cluster appears to be resistant to repeat-induced point mutations, a P450 monooxygenase gene adjacent to the cluster has been degraded by such mutations. Among the six solanapyrone cluster genes (sol1 to sol6), sol4 encodes a novel type of Zn(II)2Cys6 zinc cluster transcription factor. Deletion of sol4 resulted in the complete loss of solanapyrone production but did not compromise growth, sporulation, or virulence. Gene expression studies with the sol4 deletion and sol4-overexpressing mutants delimited the boundaries of the solanapyrone gene cluster and revealed that sol4 is likely a specific regulator of solanapyrone biosynthesis and appears to be necessary and sufficient for induction of the solanapyrone cluster genes. Despite the dynamic surrounding genomic regions, the solanapyrone gene cluster has maintained its integrity, suggesting important roles of solanapyrones in fungal biology.T he subtelomere, the region upstream of the telomere on a linear chromosome, is characterized by richness in repetitive sequences and frequent chromosomal rearrangement and lacks synteny among closely related species and even within the same species in fungi (1-4). Subtelomeric regions are also enriched with duplicate, translocated, and horizontally transferred genes and thought to be evolutionarily labile and important for niche adaptation (2, 5, 6). These plastic chromosomal regions often include contingency genes, such as genes involved in membrane transport (2, 6), nutrient assimilation (2, 7), anaerobiosis (8), and virulence (1, 9, 10). Secondary metabolite (SM) gene clusters also exhibit a subtelomeric bias (11)(12)(13)(14).In fungi, SMs play diverse roles in host and niche specialization, serving as virulence factors, antibiotics, and metal ionchelating agents (15, 16). Solanapyrones are polyketide-derived SMs that were originally found in the plant-pathogenic fungi Alternaria solani and Ascochyta rabiei (17, 18), as well as in some other ascomycetous fungi occupying different ecological niches (19-21). The fact that solanapyrones are produced by distantly related species like Nigrospora (Sordariomycetes) but not by closely related species of Ascochyta or Alternaria (Dothideomycetes) suggests the acquisition of the entire gene cluster by horizontal gene transfer (HGT) events. Recently, th...

The botrydial biosynthetic gene cluster of Botrytis cinerea displays a bipartite genomic structure and is positively regulated by the putative Zn(II)2Cys6 transcription factor BcBot6

“…Two A + T-rich regions (above 80% of A + T) are present upstream from the BOT gene cluster and between Bcbot5 and Bcbot6 genes. This particular genomic environment has already been described for the solanapyrone gene cluster in Ascochyta rabiei (Kim et al, 2015). In their study, the authors pointed out the presence of Tcl/Mariner-type TEs and further demonstrated that those sequences probably underwent the action of RIP resulting in A + T-rich regions.…”

Production of the antibiotic secondary metabolite solanapyrone A by the fungal plant pathogen Ascochyta rabiei during fruiting body formation in saprobic growth

“…Previously, we developed sol4‐ deletion (Δ sol4 ) and sol5 ‐deletion (Δ sol5 ) mutants that completely lost solanapyrone production (Kim et al ., ). To test antagonistic roles of solanapyrone A, the abilities to inhibit growth of other fungi that commonly isolated from chickpea debris were compared between A. rabiei wild‐type isolate AR628 (WT) and solanapyrone‐minus mutants in confrontation assays.…”

“…Solanapyrone A has long been considered to play a crucial role in causing disease of chickpea ( Cicer arietinum L.), due to its strong phytotoxicity to various plants (Ichihara et al ., ; Höhl et al ., ; Kaur, ; Hamid and Strange, ). However, recent studies using genetically defined solanopyrone‐minus mutants showed that solanapyrone A is not required for pathogenicity (Kim et al ., ).…”

“…The solanapyrone biosynthesis gene cluster was first identified in Al. solani (Kasahara et al, 2010), and later in A. rabiei (Kim et al, 2015a). The cluster comprises six genes (sol1-sol6) encoding all the necessary biosynthetic enzymes (Fig.…”

“…The cluster comprises six genes (sol1-sol6) encoding all the necessary biosynthetic enzymes (Fig. 1A) as well as a GAL4-type transcriptional regulator (sol4) for solanapyrone production (Kim et al, 2015a). Among the cluster genes, the sol1 gene encodes a polyketide synthase that initiates the solanapyrone biosynthesis pathway, and sol5, the first proven natural Diels-Alderase gene, catalyzes both oxidation and subsequent cyclization of the immediate precursor compound of solanapyrone A, as illustrated in Fig.…”