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We have identified the tomato I gene for resistance to the Fusarium wilt fungus Fusarium oxysporum f. sp. lycopersici ( Fol) and show that it encodes a membrane-anchored leucine-rich repeat receptor-like protein ( LRR- RLP). Unlike most other LRR- RLP genes involved in plant defence, the I gene is not a member of a gene cluster and contains introns in its coding sequence. The I gene encodes a loopout domain larger than those in most other LRR- RLPs, with a distinct composition rich in serine and threonine residues. The I protein also lacks a basic cytosolic domain. Instead, this domain is rich in aromatic residues that could form a second transmembrane domain. The I protein recognises the Fol Avr1 effector protein, but, unlike many other LRR- RLPs, recognition specificity is determined in the C-terminal half of the protein by polymorphic amino acid residues in the LRRs just preceding the loopout domain and in the loopout domain itself. Despite these differences, we show that I/Avr1-dependent necrosis in Nicotiana benthamiana depends on the LRR receptor-like kinases ( RLKs) SERK3/ BAK1 and SOBIR1. Sequence comparisons revealed that the I protein and other LRR- RLPs involved in plant defence all carry residues in their last LRR and C-terminal LRR capping domain that are conserved with SERK3/ BAK1-interacting residues in the same relative positions in the LRR- RLKs BRI1 and PSKR1. Tyrosine mutations of two of these conserved residues, Q922 and T925, abolished I/Avr1-dependent necrosis in N. benthamiana, consistent with similar mutations in BRI1 and PSKR1 preventing their interaction with SERK3/ BAK1. [ABSTRACT FROM AUTHOR]

The genomes of many filamentous fungi consist of a ‘core’ part containing conserved genes essential for normal development as well as conditionally dispensable (CD) or lineage-specific (LS) chromosomes. In the plant-pathogenic fungus Fusarium oxysporum f. sp. lycopersici, one LS chromosome harbours effector genes that contribute to pathogenicity. We employed flow cytometry to select for events of spontaneous (partial) loss of either the two smallest LS chromosomes or two different core chromosomes. We determined the rate of spontaneous loss of the ‘effector’ LS chromosome in vitro at around 1 in 35 000 spores. In addition, a viable strain was obtained lacking chromosome 12, which is considered to be a part of the core genome. We also isolated strains carrying approximately 1-Mb deletions in the LS chromosomes and in the dispensable core chromosome. The large core chromosome 1 was never observed to sustain deletions over 200 kb. Whole-genome sequencing revealed that some of the sites at which the deletions occurred were the same in several independent strains obtained for the two chromosomes tested, indicating the existence of deletion hotspots. For the core chromosome, this deletion hotspot was the site of insertion of the marker used to select for loss events. Loss of the core chromosome did not affect pathogenicity, whereas loss of the effector chromosome led to a complete loss of pathogenicity. [ABSTRACT FROM AUTHOR]

Horizontal transfer of supernumerary or lineage-specific (LS) chromosomes has been described in a number of plant pathogenic filamentous fungi. So far it was not known whether transfer is restricted to chromosomes of certain size or properties, or whether ‘core’ chromosomes can also undergo horizontal transfer. We combined a directed and a non-biased approach to determine whether such restrictions exist. Selection genes were integrated into the genome of a strain of Fusarium oxysporum pathogenic on tomato, either targeted to specific chromosomes by homologous recombination or integrated randomly into the genome. By testing these strains for transfer of the marker to another strain we could confirm transfer of a previously described mobile pathogenicity chromosome. Surprisingly, we also identified strains in which (parts of) core chromosomes were transferred. Whole genome sequencing revealed that this was accompanied by the loss of the homologous region from the recipient strain. Remarkably, transfer of the mobile pathogenicity chromosome always accompanied this exchange of core chromosomes. [ABSTRACT FROM AUTHOR]

• Development of resistant crops is the most effective way to control plant diseases to safeguard food and feed production. Disease resistance is commonly based on resistance genes, which generally mediate the recognition of small proteins secreted by invading pathogens. These proteins secreted by pathogens are called 'avirulence' proteins. Their identification is important for being able to assess the usefulness and durability of resistance genes in agricultural settings. • We have used genome sequencing of a set of strains of the melon wilt fungus Fusarium oxysporum f. sp. melonis (Fom), bioinformatics-based genome comparison and genetic transformation of the fungus to identify AVRFOM2, the gene that encodes the avirulence protein recognized by the melon Fom-2 gene. • Both an unbiased and a candidate gene approach identified a single candidate for the AVRFOM2 gene. Genetic complementation of AVRFOM2 in three different race 2 isolates resulted in resistance of Fom-2-harbouring melon cultivars. AvrFom2 is a small, secreted protein with two cysteine residues and weak similarity to secreted proteins of other fungi. • The identification of AVRFOM2 will not only be helpful to select melon cultivars to avoid melon Fusarium wilt, but also to monitor how quickly a Fom population can adapt to deployment of Fom-2-containing cultivars in the field. [ABSTRACT FROM AUTHOR]

Plant-invading microbes betray their presence to a plant by exposure of antigenic molecules such as small, secreted proteins called 'effectors'. In Fusarium oxysporum f. sp . lycopersici ( Fol) we identified a pair of effector gene candidates, AVR2- SIX5, whose expression is controlled by a shared promoter., The pathogenicity of AVR2 and SIX5 Fol knockouts was assessed on susceptible and resistant tomato ( Solanum lycopersicum) plants carrying I-2. The I-2 NB- LRR protein confers resistance to Fol races carrying AVR2., Like Avr2, Six5 was found to be required for full virulence on susceptible plants. Unexpectedly, each knockout could breach I-2-mediated disease resistance. So whereas Avr2 is sufficient to induce I-2-mediated cell death, Avr2 and Six5 are both required for resistance. Avr2 and Six5 interact in yeast two-hybrid assays as well as in planta. Six5 and Avr2 accumulate in xylem sap of plants infected with the reciprocal knockouts, showing that lack of I-2 activation is not due to a lack of Avr2 accumulation in the SIX5 mutant., The effector repertoire of a pathogen determines its host specificity and its ability to manipulate plant immunity. Our findings challenge an oversimplified interpretation of the gene-for-gene model by showing requirement of two fungal genes for immunity conferred by one resistance gene. [ABSTRACT FROM AUTHOR]

Genome sequencing of F usarium oxysporum revealed that pathogenic forms of this fungus harbour supernumerary chromosomes with a wide variety of genes, many of which likely encode traits required for pathogenicity or niche specialization. Specific transcription factor gene families are expanded on these chromosomes including the EBR1 family ( Enhanced Branching). The significance of the EBR1 family expansion on supernumerary chromosomes and whether EBR1 paralogues are functional is currently unknown. EBR1 is found as a single copy in F. graminearum and other fungi but as multiple paralogues in pathogenic F . oxysporum strains. These paralogues exhibit sequence and copy number variation among different host-specific strains and even between more closely related strains. Relative expression of the EBR1 paralogues depends on growth conditions and on the presence of the single EBR1 gene in the core genome. Deletion of EBR1 in the core genome in different F . oxysporum strains resulted in impaired growth, reduced pathogenicity and slightly reduced biocontrol capacities. To identify genes regulated by EBR1, the transcriptomes of wild-type and Δ ebr1 strains were compared for both F . oxysporum and F . graminearum. These studies showed that in both species, EBR1 regulates genes involved in general metabolism as well as virulence. [ABSTRACT FROM AUTHOR]