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Abstract The genomes of the fungus Magnaporthe oryzae that causes blast diseases on diverse grass species, including major crops, have indispensable core-chromosomes and may contain supernumerary chromosomes, also known as mini-chromosomes. These mini-chromosomes are speculated to provide effector gene mobility, and may transfer between strains. To understand the biology of mini-chromosomes, it is valuable to be able to detect whether a M. oryzae strain possesses a mini-chromosome. Here, we applied recurrent neural network models for classifying DNA sequences as arising from core- or mini-chromosomes. The models were trained with sequences from available core- and mini-chromosome assemblies, and then used to predict the presence of mini-chromosomes in a global collection of M. oryzae isolates using short-read DNA sequences. The model predicted that mini-chromosomes were prevalent in M. oryzae isolates. Interestingly, at least one mini-chromosome was present in all recent wheat isolates, but no mini-chromosomes were found in early isolates collected before 1991, indicating a preferential selection for strains carrying mini-chromosomes in recent years. The model was also used to identify assembled contigs derived from mini-chromosomes. In summary, our study has developed a reliable method for categorizing DNA sequences and showcases an application of recurrent neural networks in predictive genomics. 

Abstract Goss's wilt, caused by the Gram-positive actinobacterium Clavibacter nebraskensis, is an important bacterial disease of maize. The molecular and genetic mechanisms of resistance to the bacterium, or, in general, Gram-positive bacteria causing plant diseases, remain poorly understood. Here, we examined the genetic basis of Goss's wilt through differential gene expression, standard genome-wide association mapping (GWAS), extreme phenotype (XP) GWAS using highly resistant (R) and highly susceptible (S) lines, and quantitative trait locus (QTL) mapping using 3 bi-parental populations, identifying 11 disease association loci. Three loci were validated using near-isogenic lines or recombinant inbred lines. Our analysis indicates that Goss's wilt resistance is highly complex and major resistance genes are not commonly present. RNA sequencing of samples separately pooled from R and S lines with or without bacterial inoculation was performed, enabling identification of common and differential gene responses in R and S lines. Based on expression, in both R and S lines, the photosynthesis pathway was silenced upon infection, while stress-responsive pathways and phytohormone pathways, namely, abscisic acid, auxin, ethylene, jasmonate, and gibberellin, were markedly activated. In addition, 65 genes showed differential responses (up- or down-regulated) to infection in R and S lines. Combining genetic mapping and transcriptional data, individual candidate genes conferring Goss's wilt resistance were identified. Collectively, aspects of the genetic architecture of Goss's wilt resistance were revealed, providing foundational data for mechanistic studies. 

Summary The fungal pathogen,Magnaporthe oryzae Triticumpathotype, causing wheat blast disease was first identified in South America and recently spread across continents to South Asia and Africa. Here, we studied the genetic relationship among isolates found on the three continents.Magnaporthe oryzaestrains closely related to a South American field isolate B71 were found to have caused the wheat blast outbreaks in South Asia and Africa. Genomic variation among isolates from the three continents was examined using an improved B71 reference genome and whole‐genome sequences.We found strong evidence to support that the outbreaks in Bangladesh and Zambia were caused by the introductions of genetically separated isolates, although they were all close to B71 and, therefore, collectively referred to as the B71 branch. In addition, B71 branch strains carried at least one supernumerary mini‐chromosome. Genome assembly of a Zambian strain revealed that its mini‐chromosome was similar to the B71 mini‐chromosome but with a high level of structural variation.Our findings show that while core genomes of the multiple introductions are highly similar, the mini‐chromosomes have undergone marked diversification. The maintenance of the mini‐chromosome and rapid genomic changes suggest the mini‐chromosomes may serve important virulence or niche adaptation roles under diverse environmental conditions. 

Abstract Understanding gene regulatory networks is essential to elucidate developmental processes and environmental responses. Here, we studied regulation of a maize (Zea mays) transcription factor gene using designer transcription activator-like effectors (dTALes), which are synthetic Type III TALes of the bacterial genus Xanthomonas and serve as inducers of disease susceptibility gene transcription in host cells. The maize pathogen Xanthomonas vasicola pv. vasculorum was used to introduce 2 independent dTALes into maize cells to induced expression of the gene glossy3 (gl3), which encodes a MYB transcription factor involved in biosynthesis of cuticular wax. RNA-seq analysis of leaf samples identified, in addition to gl3, 146 genes altered in expression by the 2 dTALes. Nine of the 10 genes known to be involved in cuticular wax biosynthesis were upregulated by at least 1 of the 2 dTALes. A gene previously unknown to be associated with gl3, Zm00001d017418, which encodes aldehyde dehydrogenase, was also expressed in a dTALe-dependent manner. A chemically induced mutant and a CRISPR-Cas9 mutant of Zm00001d017418 both exhibited glossy leaf phenotypes, indicating that Zm00001d017418 is involved in biosynthesis of cuticular waxes. Bacterial protein delivery of dTALes proved to be a straightforward and practical approach for the analysis and discovery of pathway-specific genes in maize. 

Abstract DNA double-strand breaks require repair or risk corrupting the language of life. To ensure genome integrity and viability, multiple DNA double-strand break repair pathways function in eukaryotes. Two such repair pathways, canonical non-homologous end joining and homologous recombination, have been extensively studied, while other pathways such as microhomology-mediated end joint and single-strand annealing, once thought to serve as back-ups, now appear to play a fundamental role in DNA repair. Here, we review the molecular details and hierarchy of these four DNA repair pathways, and where possible, a comparison for what is known between animal and fungal models. We address the factors contributing to break repair pathway choice, and aim to explore our understanding and knowledge gaps regarding mechanisms and regulation in filamentous pathogens. We additionally discuss how DNA double-strand break repair pathways influence genome engineering results, including unexpected mutation outcomes. Finally, we review the concept of biased genome evolution in filamentous pathogens, and provide a model, termed Biased Variation, that links DNA double-strand break repair pathways with properties of genome evolution. Despite our extensive knowledge for this universal process, there remain many unanswered questions, for which the answers may improve genome engineering and our understanding of genome evolution. 

Abstract The wheat wild relativeAegilops tauschiiwas previously used to transfer theLr42leaf rust resistance gene into bread wheat.Lr42confers resistance at both seedling and adult stages, and it is broadly effective against all leaf rust races tested to date.Lr42has been used extensively in the CIMMYT international wheat breeding program with resulting cultivars deployed in several countries. Here, using a bulked segregant RNA-Seq (BSR-Seq) mapping strategy, we identify three candidate genes forLr42. Overexpression of a nucleotide-binding site leucine-rich repeat (NLR) gene AET1Gv20040300 induces strong resistance to leaf rust in wheat and a mutation of the gene disrupted the resistance. TheLr42resistance allele is rare inAe. tauschiiand likely arose from ectopic recombination. Cloning ofLr42provides diagnostic markers and over 1000 CIMMYT wheat lines carryingLr42have been developed documenting its widespread use and impact in crop improvement. 

Through the association of protein complexes to DNA, the eukaryotic nuclear genome is broadly organized into open euchromatin that is accessible for enzymes acting on DNA and condensed heterochromatin that is inaccessible. Chemical and physical alterations to chromatin may impact its organization and functionality and are therefore important regulators of nuclear processes. Studies in various fungal plant pathogens have uncovered an association between chromatin organization and expression of in planta - induced genes that are important for pathogenicity. This review discusses chromatin-based regulation mechanisms as determined in the fungal plant pathogen Verticillium dahliae and relates the importance of epigenetic transcriptional regulation and other nuclear processes more broadly in fungal plant pathogens.