Ascospores are the primary inoculum in Fusarium graminearum . Interestingly, 70 of its genes have premature stop codons (PSC) and require A-to-I editing during sexual reproduction to encode full-length proteins, including the ortholog of yeast Ama1, a meiosis-specific activator of APC/C. In this study, we characterized the function of FgAMA1 and its PSC editing. FgAMA1 was specifically expressed during sexual reproduction. The Fgama1 mutant was normal in growth and perithecium formation but defective in ascospogenesis. Instead of forming four-celled, uninucleate ascospores, Fgama1 mutant produced oval, single-celled, binucleated ascospores by selfing. Some mutant ascospores began to bud and underwent additional mitosis inside asci. Expression of the wild-type or edited FgAMA1 but not the uneditable allele complemented Fgama1 . In the Fgama1 x mat-1-1 outcross, over 60% of the asci had eight Fgama1 or intermediate (elongated but single-celled) ascospores, suggesting efficient meiotic silencing of unpaired FgAMA1 . Deletion of FgPAL1 , one of the genes upregulated in Fgama1 also resulted in defects in ascospore morphology and budding. Overall, our results showed that FgAMA1 is dispensable for meiosis but important for ascospore formation and discharge. In F. graminearum , whereas some of its targets are functional during meiosis, FgAma1 may target other proteins that function after spore delimitation.

The basal ascomycetes in genus Taphrina have strict host specificity and coevolution with their host plants, making them appealing models for studying the genomic basis of ecological divergence and host adaption. We therefore performed genome sequencing and comparative genomics of different Taphrina species with distinct host ranges to reveal their evolution. We identified frequent chromosomal rearrangements and highly dynamic lineage‐specific (LS) genomic regions in Taphrina genomes. The LS regions occur at the flanking regions of chromosomal breakpoints, and are greatly enriched for DNA repeats, non‐core genes, and in planta up‐regulated genes. Furthermore, we identified hundreds of candidate secreted effector proteins (CSEPs) that are commonly organized in gene clusters that form distinct AT‐rich isochore‐like regions. Nearly half of the CSEPs constitute two novel superfamilies with modular structures unique to Taphrina. These CSEPs are commonly up‐regulated during infection, enriched in the LS regions, evolved faster, and underwent extensive gene gain and loss in different species. In addition to displaying signatures of positive selection, functional characterization of selected CSEP genes confirmed their roles in suppression of plant defence responses. Overall, our results showed that extensive chromosomal rearrangements and rapidly evolving CSEP superfamilies play important roles in speciation and host adaptation in the early‐branching ascomycetous fungi.

The versatile functions of SR (serine/arginine-rich) proteins in pre-mRNA splicing and processing are modulated by reversible phosphorylation. Previous studies showed that FgPrp4, the only protein kinase among spliceosome components, is important for intron splicing and the FgSrp1 SR protein is phosphorylated at five conserved sites in Fusarium graminearum. In this study, we showed that the Fgsrp1 deletion mutant rarely produced conidia and caused only limited symptoms on wheat heads and corn silks. Deletion of FgSRP1 also reduced ascospore ejection and deoxynivalenol (DON) production. Interestingly, FgSRP1 had two transcript isoforms due to alternative splicing and both of them were required for its normal functions in growth and DON biosynthesis. FgSrp1 localized to the nucleus and interacted with FgPrp4 in vivo. Deletion of all four conserved phosphorylation sites but not individual ones affected the FgSRP1 function, suggesting their overlapping functions. RNA-seq analysis showed that the expression of over thousands of genes and splicing efficiency in over 140 introns were affected. Taken together, FgSRP1 is important for conidiation, and pathogenesis and alternative splicing is important for its normal functions. The FgSrp1 SR protein is likely important for pre-mRNA processing or splicing of various genes in different developmental and infection processes.

Fusarium graminearum is a causal agent of wheat scab disease and a producer of deoxynivalenol (DON) mycotoxins. Treatment with exogenous cyclic adenosine monophosphate (cAMP) increases its DON production. In this study, to better understand the role of the cAMP-protein kinase A (PKA) pathway in F. graminearum, we functionally characterized the PKR gene encoding the regulatory subunit of PKA. Mutants deleted of PKR were viable, but showed severe defects in growth, conidiation and plant infection. The pkr mutant produced compact colonies with shorter aerial hyphae with an increased number of nuclei in hyphal compartments. Mutant conidia were morphologically abnormal and appeared to undergo rapid autophagy-related cell death. The pkr mutant showed blocked perithecium development, but increased DON production. It had a disease index of less than unity and failed to spread to neighbouring spikelets. The mutant was unstable and spontaneous suppressors with a faster growth rate were often produced on older cultures. A total of 67 suppressor strains that grew faster than the original mutant were isolated. Three showed a similar growth rate and colony morphology to the wild-type, but were still defective in conidiation. Sequencing analysis with 18 candidate PKA-related genes in three representative suppressor strains identified mutations only in the CPK1 catalytic subunit gene. Further characterization showed that 10 of the other 64 suppressor strains also had mutations in CPK1. Overall, these results showed that PKR is important for the regulation of hyphal growth, reproduction, pathogenesis and DON production, and mutations in CPK1 are partially suppressive to the deletion of PKR in F. graminearum.

Summary Ascospores generated during sexual reproduction are the primary inoculum for the wheat scab fungus Fusarium graminearum. Purine metabolism is known to play important roles in fungal pathogens but its lifecycle stage‐specific regulation is unclear. By characterizing the genes involved in purine de novo and salvage biosynthesis pathways, we showed that de novo syntheses of inosine, adenosine and guanosine monophosphates (IMP, AMP and GMP) are important for vegetative growth, sexual/asexual reproduction, and infectious growth, whereas purine salvage synthesis is dispensable for these stages in F. graminearum. Addition of GMP rescued the defects of the Fgimd1 mutant in vegetative growth and conidiation but not sexual reproduction, whereas addition of AMP rescued all of these defects of the Fgade12 mutant, suggesting that the function of de novo synthesis of GMP rather than AMP is distinct in sexual stages. Moreover, Acd1, an ortholog of AMP deaminase, is dispensable for growth but essential for ascosporogenesis and pathogenesis, suggesting that AMP catabolism has stage‐specific functions during sexual reproduction and infectious growth. The expression of almost all the genes involved in de novo purine synthesis is downregulated during sexual reproduction and infectious growth relative to vegetative growth. This study revealed that F. graminearum has stage‐specific regulation of purine metabolism during infectious growth and sexual reproduction.