Although fungi lack adenosine deaminase acting on RNA (ADAR) enzymes, adenosine to inosine (A-to-I) RNA editing was reported recently in during sexual reproduction. In this study, we profiled the A-to-I editing landscape and characterized its functional and adaptive properties in the model filamentous fungus A total of 40,677 A-to-I editing sites were identified, and approximately half of them displayed stage-specific editing or editing levels at different sexual stages. RNA-sequencing analysis with the Δ-1 and Δ-1 mutants confirmed A-to-I editing occurred before ascus development but became more prevalent during ascosporogenesis. Besides fungal-specific sequence and secondary structure preference, 63.5% of A-to-I editing sites were in the coding regions and 81.3% of them resulted in nonsynonymous recoding, resulting in a significant increase in the proteome complexity. Many genes involved in RNA silencing, DNA methylation, and histone modifications had extensive recoding, including -1,-3, -1, and-2. Fifty pseudogenes harbor premature stop codons that require A-to-I editing to encode full-length proteins. Unlike in humans, nonsynonymous editing events in are generally beneficial and favored by positive selection. Almost half of the nonsynonymous editing sites in are conserved and edited in Furthermore, hundreds of them are conserved in and had higher editing levels. Two unknown genes with editing sites conserved between and were experimentally shown to be important for ascosporogenesis. This study comprehensively analyzed A-to-I editing in and showed that RNA editing is stage-specific and generally adaptive, and may be functionally related to repeat induced point mutation and meiotic silencing by unpaired DNA.

Fusarium head blight caused by Fusarium graminearum is an important disease of wheat and barley worldwide. In a previous study on functional characterization of the F. graminearum kinome, one protein kinase gene important for virulence is orthologous to SCH9 that is functionally related to the cAMP-PKA and TOR pathways in the budding yeast. In this study, we further characterized the functions of FgSCH9 in F. graminearum and its ortholog in Magnaporthe oryzae. The ΔFgsch9 mutant was slightly reduced in growth rate but significantly reduced in conidiation, DON production, and virulence on wheat heads and corn silks. It had increased tolerance to elevated temperatures but became hypersensitive to oxidative, hyperosmotic, cell wall, and membrane stresses. The ΔFgsch9 deletion also had conidium morphology defects and produced smaller conidia. These results suggest that FgSCH9 is important for stress responses, DON production, conidiogenesis, and pathogenesis in F. graminearum. In the rice blast fungus Magnaporthe oryzae, the ΔMosch9 mutant also was defective in conidiogenesis and pathogenesis. Interestingly, it also produced smaller conidia and appressoria. Taken together, our data indicate that the SCH9 kinase gene may have a conserved role in regulating conidium size and plant infection in phytopathogenic ascomycetes.

No abstract

Trichothecene mycotoxins, such as deoxynivalenol (DON) produced by the fungal pathogen, Fusarium graminearum, are not only important for plant infection but are also harmful to human and animal health. Trichothecene targets the ribosomal protein Rpl3 that is conserved in eukaryotes. Hence, a self-defense mechanism must exist in DON-producing fungi. It is reported that TRI (trichothecene biosynthesis) 101 and TRI12 are two genes responsible for self-defense against trichothecene toxins in Fusarium. In this study, however, we found that simultaneous disruption of TRI101 and TRI12 has no obvious influence on DON resistance upon exogenous DON treatment in F. graminearum, suggesting that other mechanisms may be involved in self-defense. By using RNA-seq, we identified 253 genes specifically induced in DON-treated cultures compared with samples from cultures treated or untreated with cycloheximide, a commonly used inhibitor of eukaryotic protein synthesis. We found that transporter genes are significantly enriched in this group of DON-induced genes. Of those genes, 15 encode major facilitator superfamily transporters likely involved in mycotoxin efflux. Significantly, we found that genes involved in the metabolism of gamma-aminobutyric acid (GABA), a known inducer of DON production in F. graminearum, are significantly enriched among the DON-induced genes. The GABA biosynthesis gene PROLINE UTILIZATION 2-2 (PUT2-2) is downregulated, while GABA degradation genes are upregulated at least twofold upon treatment with DON, resulting in decreased levels of GABA. Taken together, our results suggest that transporters influencing DON efflux are important for self-defense and that GABA mediates the balance of DON production and self-defense in F. graminearum.

Summary Alternative splicing (AS) and alternative polyadenylation (APA) contribute significantly to the regulation of gene expression in higher eukaryotes. Their biological impact in filamentous fungi, however, is largely unknown. Here we combine PacBio Isoform‐Sequencing and strand‐specific RNA‐sequencing of multiple tissues and mutant characterization to reveal the landscape and regulation of AS and APA in Fusarium graminearum. We generated a transcript annotation comprising 51 617 isoforms from 17 189 genes. In total, 4997 and 11 133 genes are alternatively spliced and polyadenylated, respectively. Majority of the AS events alter coding sequences. Unexpectedly, the AS transcripts containing premature‐termination codons are not sensitive to nonsense‐mediated messenger RNA decay. Unlike in yeasts and animals, distal APA sites have strong signals, but proximal APA isoforms are highly expressed in F. graminearum. The 3′‐end processing factors FgRNA15, FgHRP1, and FgFIP1 play roles in promoting proximal APA site usage and intron splicing. A genome‐wide increase in intron inclusion and distal APA site usage and downregulation of the spliceosomal and 3′‐end processing factors were observed in older and quiescent tissues, indicating intron inclusion and 3′‐untranslated region lengthening as novel mechanisms in regulating aging and dormancy in fungi. This study provides new insights into the complexity and regulation of AS and APA in filamentous fungi.

Summary The wheat head blight fungus Fusarium graminearum has two highly similar beta‐tubulin genes with overlapping functions during vegetative growth but only TUB1 is important for sexual reproduction. To better understand their functional divergence during ascosporogenesis, in this study we characterized the sequence elements important for stage‐specific functions of TUB1. Deletion of TUB1 blocked the late but not initial stages of perithecium formation. Perithecia formed by tub1 mutant had limited ascogenous hyphae and failed to develop asci. Silencing of TUB1 by MSUD also resulted in defects in ascospore formation. Interestingly, the 3′‐UTR of TUB1 was dispensable for growth but essential for its function during sexual reproduction. RIP mutations that specifically affected Tub1 functions during sexual reproduction also were identified in two ascospore progeny. Furthermore, site‐directed mutagenesis showed that whereas the non‐editable mutations at three A‐to‐I RNA editing sites had no effects, the N347D (not T362D or I368V) edited mutation affected ascospore development. In addition, the F167Y, but not E198K or F200Y, mutation in TUB1 conferred tolerance to carbendazim and caused a minor defect in sexual reproduction. Taken together, our data indicate TUB1 plays an essential role in ascosporogenesis and sexual‐specific functions of TUB1 require stage‐specific RNA processing and Tub1 expression.

Here, we report a novel mycotombus-like mycovirus, tentatively named 'Phoma matteucciicola RNA virus 2' (PmRV2), derived from the phytopathogenic fungus Phoma matteucciicola strain HNQH1. The complete PmRV2 genome is comprised of 3,460 nucleotides (nts), a positive-sense single-stranded RNA (+ ssRNA) with a GC content of 56.71%. Sequence analysis of PmRV2 indicated the presence of two noncontiguous open reading frames (ORFs) encoding a hypothetical protein and a RNA-dependent RNA polymerase (RdRp), respectively. PmRV2 contains a metal-binding 'GDN' triad in Motif C of RdRp while most + ssRNA mycoviruses contained a 'GDD' motif in the same region. Additionally, a BLASTp search showed that the RdRp amino acid sequence of PmRV2 was most closely related to the RdRp of Macrophomina phaseolina umbra-like virus 1 (50.72% identity) and Erysiphe necator umbra-like virus 2 (EnUlV2, 44.84% identity). Phylogenetic analysis indicated that PmRV2 grouped together with EnUlV2 within the recently proposed family of 'Mycotombusviridae'.

No abstract