Analysis of intron positions in genes with recent insertions of Introner-Like Elements (ILE).

<p>ILEs from <i>Passalora brachycarpa</i> inserted in genes encoding (A) a transporter and (B) a peroxidase. ILEs from <i>Passalora miurae</i> inserted in genes encoding (C) a hydroxylase/oxidoreductase and (D) a fungal transcription factor. For each gene, a maximum likelihood phylogenetic tree was constructed with the predicted protein sequence of orthologues. The trees were rooted with the closest homologue found in Basidiomycota. Bootstrap values of 100 repeats are shown. Scale bar represents the number of substitutions per site. The numbers correspond to the protein ID from the Joint Genome Institute mycocosm portal, except for one gene that was not predicted in the <i>Fulvia fulva</i> genome and for which genomic coordinates are given (B). Orders in fungal classification are mentioned in the trees. On the right, diagrams depict aligned protein sequences and intron positions are indicated as black bars. Their positions in the protein alignment that served to build the phylogenetic trees are indicated above. Positions that are shown in grey highlight putative intron sliding. The black arrows indicate the positions where ILEs inserted in the genes of <i>P</i>. <i>miurae</i> or <i>P</i>. <i>brachycarpa</i>. The open triangles indicate previously identified ILEs. Dots indicate positions where parallel intron gains have occurred. The black bar below each protein representation indicates conserved domains (positions in the protein alignments are indicated between brackets). Asterisks behind species names indicate genes that are likely pseudogenes because of an in frame stop codon. (E) Schematic overview of intron positions (numbered on top) in three genes. The first row shows intron positions in genes of <i>P</i>. <i>miurae</i> or <i>P</i>. <i>brachycarpa</i>. Thick lines indicate monophyletic clades according to the phylogenetic trees. Black, dark grey and light grey squares indicate single presence of an intron position in a monophyletic clade, presence-absence polymorphism and single absence of an intron position in a monophyletic clade, respectively. White squares indicate absence of intron. The presence of ILE and occurrence of putative intron splicing (IS) are indicated below each scheme. <i>Aciri</i>: <i>Acidomyces richmondensis</i>; <i>Altbr</i>: <i>Alternaria brassicicola</i>; <i>Cerzm</i>: <i>Cercospora zeae-maydis</i>; <i>Fulfu</i>: <i>Fulvia fulva</i>; <i>Bipze</i>: <i>Bipolaris zeicola</i>; <i>Bipma</i>: <i>Bipolaris maydis</i>; <i>Curlu</i>: <i>Curvularia lunata</i>; <i>Bipor</i>: <i>Bipolaris oryzae</i>; <i>Bipso</i>: <i>Bipolaris sorokiniana</i>; <i>Bipvi</i>: <i>Bipolaris victoriae</i>; <i>Didex</i>: <i>Didymella exigua</i>; <i>Dotse</i>: <i>Dothistroma septosporum</i>; <i>Lentfl</i>: <i>Lentithecium fluviatile</i>; <i>Pleli</i>: <i>Plenodomus lingam</i>; <i>Lopma</i>: <i>Lophiostoma macrostomum</i>; <i>Psefi</i>: <i>Pseudocercospora fijiensis</i>; <i>Pyrtr</i>: <i>Pyrenophora tritici-repentis</i>; <i>Pyrtt</i>: <i>Pyrenophora teres f</i>. <i>teres</i>; <i>Sphmu</i>: <i>Sphaerulina musiva</i>; <i>Sphpo</i>: <i>Sphaerulina populicola</i>; <i>Parno</i>: <i>Parastagonospora nodorum</i>; <i>Zasce</i>: <i>Zasmidium cellare</i>; <i>Zymtr</i>: <i>Zymoseptoria tritici</i>; <i>Aspve</i>: <i>Aspergillus versicolor</i>; <i>Aspwe</i>: <i>Aspergillus wentii</i>; <i>Penbi</i>: <i>Penicillium bilaiae</i>; <i>Penbr</i>: <i>Penicillium brevicompactum</i>; <i>Penca</i>: <i>Penicillium canescens</i>; <i>Pench</i>: <i>Penicillium chrysogenum</i>; <i>Penfe</i>: <i>Penicillium fellutanum</i>; <i>Pengl</i>: <i>Penicillium glabrum</i>; <i>Penox</i>: <i>Penicillium oxalicum</i>; <i>Colgr</i>: <i>Colletotrichum graminicola</i>; <i>Colfi</i>: <i>Colletotrichum fiorinae</i>; <i>Colgl</i>: <i>Colletotrichum gloeosporioides</i>; <i>Melva</i>: <i>Meliniomyces variabilis</i>; <i>Glotr</i>: <i>Gloeophyllum trabeum</i>; <i>Phlgi</i>: <i>Phlebiopsis gigantean</i>; <i>Psean</i>: <i>Pseudozyma antarctica</i>; <i>Psehu</i>: <i>Pseudozyma hubeiensis</i>.</p

Detection of Introner-Like Elements (ILEs) in closely related dothideomycetous fungal species.

<p>(A) Maximum-likelihood phylogenetic tree using ITS and LSU sequences. Fungal species in which ILEs have been previously identified are highlighted in bold. <i>Aspergillus niger</i> belongs to the <i>Eurotiomycetes</i> and serves to root the tree. Accession numbers of fungal species from the CBS-KNAW collection are indicated in between brackets. The scale bar indicates the number of substitutions per site. (B) PCR with primers specific to single (<i>cf01</i> and <i>cf02</i>) or shared (<i>cf02cf03ds01ds05</i>, <i>cf04ds03</i> and <i>cf08ds04</i>) ILE families between <i>F</i>. <i>fulva</i> and <i>Dothistroma septosporum</i> was performed using genomic DNA. PCR products were run on 20% acrylamide gels. The first row shows a 50 bp-step DNA ladder and the last row shows the water control. Asterisks indicate fragments that correspond to two different ILEs as revealed by sequencing.</p

Alignments of newly discovered Introner-Like Elements (ILEs).

<p>DNA sequences obtained from the PCR fragments were aligned, using as references the consensus sequence and the four most conserved ILE sequences of the corresponding families in <i>F</i>. <i>fulva</i> and <i>Dothistroma septosporum</i>. The complete sequences of known ILEs amplified by PCR in this study are also included. Alignments are shown for ILEs related to (A) <i>cf01</i>, (B) <i>cf02cf03ds01ds05</i> and (C) <i>cf08ds04</i> families. Bars above the alignments indicate the oligonucleotide sequences. <i>Fulful</i>: <i>Fulvia fulva</i>; <i>Dotsep</i>: <i>Dothistroma septosporum</i>; <i>Amyafr</i>: <i>Amycosphaerella africana</i>; <i>Pasbra</i>: <i>Passalora brachycarpa</i>; <i>Pascap</i>: <i>Passalora capsicicola</i>; <i>Pasdal</i>: <i>Passalora daleae</i>; <i>Pasmic</i>: <i>Passalora microsora</i>; <i>Pasmiu</i>: <i>Passalora miurae</i>; <i>Pasper</i>: <i>Passalora perfoliati</i>; <i>Passmi</i>: <i>Passalora smilacis</i>.</p

Correction: The Genomes of the Fungal Plant Pathogens <i>Cladosporium fulvum</i> and <i>Dothistroma septosporum</i> Reveal Adaptation to Different Hosts and Lifestyles But Also Signatures of Common Ancestry

<p>Correction: The Genomes of the Fungal Plant Pathogens <i>Cladosporium fulvum</i> and <i>Dothistroma septosporum</i> Reveal Adaptation to Different Hosts and Lifestyles But Also Signatures of Common Ancestry</p

Arrangement of predicted dothistromin genes in <i>Dothistroma septosporum</i> and <i>Cladosporium fulvum</i>.

<p>A) Predicted dothistromin genes within the labeled clusters (left to right) are: <i>Ver1, DotC</i> (<i>Ver1</i> cluster); <i>PksA, CypX, AvfA, MoxY</i> (<i>PksA</i> cluster); <i>AflR, AflJ</i> (<i>AflR/J</i> cluster); <i>OrdB</i>, <i>AvnA, HexB, HexA, HypC, VbsA</i> (<i>VbsA</i> cluster); <i>Nor1, AdhA, VerB</i> (<i>Nor1</i> cluster). Positions of mini-clusters are approximate and they are not drawn to scale. Dothistromin genes within the published <i>D. septosporum PksA</i> and <i>VbsA</i> clusters <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003088#pgen.1003088-Bradshaw3" target="_blank">[36]</a>, <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003088#pgen.1003088-Zhang1" target="_blank">[38]</a> and the newly discovered <i>AflR/J</i> and <i>Nor-1</i> clusters are found in the same order and orientation in <i>C. fulvum</i>. B) Expression of dothistromin biosynthetic genes (<i>Ver1, PksA, VbsA</i>) and regulatory gene (<i>AflR</i>) was determined in <i>D. septosporum</i> by quantitative PCR. Mean expression and standard deviations are shown for at least 3 biological replicates relative to β-tubulin expression. In <i>D. septosporum</i> all genes but <i>DsVbsA</i> are expressed more highly <i>in planta</i> (late-stage sporulating lesions from a forest sample) than in culture (PDB or B5 media) as highlighted by the dashed-grey line. C) Expression of <i>C. fulvum</i> genes is shown as for (B), revealing that expression is not higher during tomato infection than in culture (dashed-grey line). Note the different scales for expression, which reveal a much lower level of transcription both <i>in planta</i> and in PDB medium compared to <i>D. septosporum</i>.</p

Repetitive regions flanking known effectors of <i>Cladosporium fulvum</i>.

<p>Scaffolds harboring sequenced <i>C. fulvum</i> effector genes with flanking repeats. Repeat regions longer than 200-bp are shown, with different types indicated in different color code. Red arrows depict the effector genes and sizes are in kb. The sizes of scaffolds range from 8 to 213-kb but some are shortened to fit the figure due to differences in size.</p

Syntenic and non-syntenic regions between <i>C. fulvum</i> and <i>D. septosporum</i> are unevenly distributed over the <i>C. fulvum</i> scaffolds.

a<p>Number of repeat regions on syntenic vs. non-syntenic scaffolds.</p>b<p>A syntenic scaffold is one that contains at least a single syntenic block, but may not be syntenic along its entire length. Total syntenic scaffold size (37.4-Mb) is therefore larger than total syntenic size in whole genome (22.3-Mb).</p>c<p>Summed repeat length on syntenic <i>versus</i> non-syntenic scaffolds.</p

Occurrence of Repeat-Induced Point Mutation (RIP) signatures and repeats in <i>C. fulvum</i> and <i>D. septosporum</i>.

a<p>Number of loci, defined as consecutive blocks of sequence assigned as repeats or RIP'd regions.</p>b<p>Percentage of the genome is indicated in brackets.</p>c<p>Only classified repeats (as shown in <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003088#pgen-1003088-t002" target="_blank">Table 2</a>) were considered.</p>d<p>Repeated sequence ≥500 nt that (at least partially) overlaps with RIP'd sequence.</p>e<p>Percentage of classified repeats is indicated in brackets.</p

Organization of repeats and pathogenicity-related genes in the <i>Dothistroma septosporum</i> genome.

<p>The fourteen chromosomes from the <i>D. septosporum</i> genome assembly are shown as GC (dark grey line) and AT (pale grey line) content (%) plots made from a 500-bp sliding window using Geneious (<a href="http://www.geneious.com" target="_blank">www.geneious.com</a>). All chromosomes have telomere sequence at both ends except chromosomes 2, 11 and 14 which have telomere sequences only at the left end as shown in the figure. Chromosome 1 has been split into two parts in the figure (L, R) because of its length, and the GC/AT content scale is shown beside the right arm of this chromosome. The positions of putative <i>Avr</i> and <i>Ecp</i> effector, secondary metabolite, dothistromin biosynthesis, and mating type genes are shown above the GC/AT content plot, while the positions of repeats (>200-bp) are shown below the plot. Color-coding of the gene and repeat types is indicated in the legend. Most chromosomes have repeat clusters at one or two sites that coincide with regions of high AT content. The chromosome sizes are to scale, as indicated by the vertical pale grey lines, with the values (in kb) shown at the bottom; neither the genes nor the repeats are drawn to scale.</p

Key secondary metabolism genes in Ascomycete genomes.

<p>Numbers of predicted polyketide synthase (PKS), non-ribosomal peptide synthetase (NRPS), hybrid PKS-NRPS (Hybrid), terpene cyclase (TC) and dimethylallyl tryptophan synthase (DMATS) genes are shown. Data are from this study and from Collemare et al. 2008 <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003088#pgen.1003088-Collemare2" target="_blank">[105]</a>.</p