Åke Olson

The Norway spruce genome sequence and conifer genome evolution

Conifers have dominated forests for more than 200 million years and are of huge ecological and economic importance. Here we present the draft assembly of the 20-gigabase genome of Norway spruce (Picea abies), the first available for any gymnosperm. The number of well-supported genes (28,354) is similar to the >100 times smaller genome of Arabidopsis thaliana, and there is no evidence of a recent whole-genome duplication in the gymnosperm lineage. Instead, the large genome size seems to result from the slow and steady accumulation of a diverse set of long-terminal repeat transposable elements, possibly owing to the lack of an efficient elimination mechanism. Comparative sequencing of Pinus sylvestris, Abies sibirica, Juniperus communis, Taxus baccata and Gnetum gnemon reveals that the transposable element diversity is shared among extant conifers. Expression of 24-nucleotide small RNAs, previously implicated in transposable element silencing, is tissue-specific and much lower than in other plants. We further identify numerous long (>10,000 base pairs) introns, gene-like fragments, uncharacterized long non-coding RNAs and short RNAs. This opens up new genomic avenues for conifer forestry and breeding.

Characterization and differential expression of dhn/lea/rab-like genes during cold acclimation and drought stress in Arabidopsis thaliana

We have characterized cDNAs for two new dhn/lea/rab (dehydrin, late embryogenesis-abundant, responsive to ABA)-related genes from Arabidopsis thaliana. The two genes were strongly induced in plants exposed to low temperature (4 °C) and were accordingly designated lti45 and lti30 (low temperature-induced). The lti45 gene product contains the conserved serine stretch and three lysine-rich repeats characteristic of DHN/LEA/RAB proteins and is very similar to another low temperature-responsive protein of A. thaliana, COR47 [17]. Both proteins have the same repeat structure and an overall amino acid identity of 64%. This structural similarity of the proteins and the tandem array of the genes suggest that this gene pair arose through a duplication. The other polypeptide, LTI30, consists of several lysine-rich repeats, a structure found in CAP85, a low temperature-and water stress-responsive protein in spinach [41] and similar proteins found in wheat [20].The expression pattern of the five dhn/lea/rab-related genes (cor47, dhnX, lti30, lti45 and rab18) identified so far in A. thaliana, was characterized in plants exposed to low temperature, drought and abscisic acid (ABA). Expression of both lti30 and lti45 was mainly responsive to low temperature similar to cor47. The lti45 and lti30 genes show only a weak response to ABA in contrast to cor47, which is moderately induced by this hormone. The three genes were also induced in severely water-stressed plants although the expression of lti30 and lti45 was rather low. In contrast to these mainly low temperature-induced genes, the expression of rab18 was strongly induced both in water-stressed and ABA-treated plants but was only slightly responsive to cold. The dhnX gene showed a very different expression pattern. It was not induced with any of the treatments tested but exhibited a significant constitutive expression. The low-temperature induction of the genes in the first group, lti30 and lti45, is ABA-independent, deduced from experiments with the ABA-deficient (aba-1) and ABA-insensitive (abi1) mutants of A. thaliana, whereas the induction of rab18 is ABA-mediated. The expression of dhnX was not significantly affected in the ABA mutants.

Insight into trade???off between wood decay and parasitism from the genome of a fungal forest pathogen

Parasitism and saprotrophic wood decay are two fungal strategies fundamental for succession and nutrient cycling in forest ecosystems. An opportunity to assess the trade-off between these strategies is provided by the forest pathogen and wood decayer Heterobasidion annosum sensu lato. We report the annotated genome sequence and transcript profiling, as well as the quantitative trait loci mapping, of one member of the species complex: H. irregulare. Quantitative trait loci critical for pathogenicity, and rich in transposable elements, orphan and secreted genes, were identified. A wide range of cellulose-degrading enzymes are expressed during wood decay. By contrast, pathogenic interaction between H. irregulare and pine engages fewer carbohydrate-active enzymes, but involves an increase in pectinolytic enzymes, transcription modules for oxidative stress and secondary metabolite production. Our results show a trade-off in terms of constrained carbohydrate decomposition and membrane transport capacity during interaction with living hosts. Our findings establish that saprotrophic wood decay and necrotrophic parasitism involve two distinct, yet overlapping, processes.

Ectomycorrhizal Cortinarius species participate in enzymatic oxidation of humus in northern forest ecosystems.

In northern forests, belowground sequestration of nitrogen (N) in complex organic pools restricts nutrient availability to plants. Oxidative extracellular enzymes produced by ectomycorrhizal fungi may aid plant N acquisition by providing access to N in macromolecular complexes. We test the hypotheses that ectomycorrhizal Cortinarius species produce Mn-dependent peroxidases, and that the activity of these enzymes declines at elevated concentrations of inorganic N. In a boreal pine forest and a sub-arctic birch forest, Cortinarius DNA was assessed by 454-sequencing of ITS amplicons and related to Mn-peroxidase activity in humus samples with- and without previous N amendment. Transcription of Cortinarius Mn-peroxidase genes was investigated in field samples. Phylogenetic analyses of Cortinarius peroxidase amplicons and genome sequences were performed. We found a significant co-localization of high peroxidase activity and DNA from Cortinarius species. Peroxidase activity was reduced by high ammonium concentrations. Amplification of mRNA sequences indicated transcription of Cortinarius Mn-peroxidase genes under field conditions. The Cortinarius glaucopus genome encodes 11 peroxidases - a number comparable to many white-rot wood decomposers. These results support the hypothesis that some ectomycorrhizal fungi--Cortinarius species in particular--may play an important role in decomposition of complex organic matter, linked to their mobilization of organically bound N.

Mitochondrial control of fungal hybrid virulence

Hybrid species of fungal pathogens that infect wild and cultivated plants are emerging with new virulence and host ranges, posing a threat to agriculture and forestry. Here we show that the virulence of hybrid species of the basidiomycete fungus Heterobasidion annosum (Fr) Bref, a causal agent of root and butt rot in conifers and one of the most economically important forest pathogens, is controlled by their mitochondrial genome. Our results indicate that cooperation between organelles that contain genetic information may influence the phenotype of hybrid phytopathogens.

Expressed sequences from the basidiomycetous tree pathogen Heterobasidion annosum during early infection of scots pine

The pattern of gene expression of the basidiomycete Heterobasidion annosum, causal agent of the root rot of conifers, was analysed during its interaction with pine roots. A complementary DNA (cDNA) library was constructed from total RNA extracted from H. annosum mycelia challenged with Scots pine seedling roots for 6 and 72h. Single pass sequencing of 1148 randomly selected cDNA clones resulted in 923 expressed sequence tags (ESTs). Contig analysis and sequence comparisons identified 318 unigene sequences, of which 62 were repeatedly sampled. A putative cellular function was assigned to 223 contigs (70%) that showed a moderate to high homology to protein sequences from public databases. Variations in expression levels during the infection process were monitored on a set of 96 unigenes by reverse northern using dot hybridisation. Seven unigenes (7%) were shown to be either up (4) or down (3) regulated during interaction of the fungus with pine roots. Fungal genes differentially expressed during contact with roots include genes encoding mitochondrial proteins, a cytochrome P450 and a vacuolar ATP synthase.

Structure and organization of two closely related low-temperature-induced dhn/lea/rab-like genes in Arabidopsis thaliana L. Heynh

We have isolated a 7 kb EcoRI genomic fragment from Arabidopsis thaliana which contains, in a tandem arrangement, two closely related dhn/lea/rab-like genes, lti29 (formerly named lti45) and cor47, corresponding to previously isolated cDNA clones. Both transcripts have been shown to accumulate in response to low temperature (LT), abscisic acid (ABA) and dehydration. Alignment of the amino acid sequences of the deduced polypeptides showed that they are 67% identical. The calculated molecular masses of the two polypeptides were 29 kDa for LTI29 and 30 kDa for COR47. Both polypeptides contain one conserved serine-stretch and three lysine-rich repeats characteristic of DHN/LEA/RAB-like proteins. In addition, both LTI29 and COR47 harbour and N-terminal acidic repeat only found in a few members amongst the DHN/LEA/RAB proteins. The close distance between the two genes (separated by 2.7 kb) and their tandem organization in the A. thaliana genome as well as the overall homology at the nucleotide sequence level of the coding region suggest that the two genes have evolved through a duplication event. This seems to be a common feature among A. thaliana LT-reponsive genes.

Evolutionary history of the conifer root rot fungus Heterobasidion annosum sensu lato

We investigated two hypotheses for the origin of the root rot fungus Heterobasidion annosum species complex: (i) that geology has been an important factor for the speciation (ii) that co‐evolutionary processes with the hosts drove the divergence of the pathogen species. The H. annosum species complex consists of five species: three occur in Europe, H. annosum s.s., Heterobasidion parviporum and Heterobasidion abietinum, and two in North America, Heterobasidion irregulare and Heterobasidion occidentale; all with different but partially overlapping host preferences. The evolution of the H. annosum species complex was studied using six partially sequenced genes, between 10 and 30 individuals of each species were analysed. Neighbour‐joining trees were constructed for each gene, and a Bayesian tree was built for the combined data set. In addition, haplotype networks were constructed to illustrate the species relationships. For three of the genes, H. parviporum and H. abietinum share haplotypes supporting recent divergence and/or possible gene flow. We propose that the H. annosum species complex originated in Laurasia and that the H. annosum s.s./H. irregulare and H. parviporum/H. abietinum/H. occidentale ancestral species emerged between 45 and 60 Ma in the Palaearctic, well after the radiation of the host genera. Our data imply that H. irregulare and H. occidentale were colonizing North America via different routes. In conclusion, plate tectonics are likely to have been the main factor influencing Heterobasidion speciation and biogeography.

Pathogenic fungal species hybrids infecting plants

In this review, we give a summary of recent examples of hybridisation in fungi. Reports on hybridisation between pathogenic fungi infecting plants have increased during the last decade. Hybrids of plant pathogens may create devastating new diseases on both cultivated and wild plants; at the same time, hybridisation is one of the natural processes in the course of evolution of new taxa.

Evolutionary significance of imbalanced nuclear ratios within heterokaryons of the basidiomycete fungus Heterobasidion parviporum

Abstract Many fungi have heterokaryotic life stages in which genetically different nuclei inhabit the same cell. In basidiomycetes, the heterokaryon is the product of mating and represents a genomic union very similar to a diploid thallus, yet the maintenance of unfused nuclei suggests a more complex association of the two genomes relative to diploidy. In species with variable numbers of nuclei per heterokaryotic cell, nuclear ratios within a mycelium may possibly become imbalanced (differ from 1:1) due to nuclear competition. In this study, heterokaryons of the basidiomycete Heterobasidion parviporum were examined to determine the effects of genotype and environment on nuclear ratios within vegetative mycelia. The data reveal that nuclear ratios are frequently imbalanced, generally stable over time, and genetically determined. The nuclear ratios were affected by environment, but the observed nuclear ratios did not follow the expectations of strong selection acting on a population of nuclei. Instead, these ratios were largely driven by genetic effects and epigenetic effects. Finally, the data suggest that nuclear ratio imbalance also affects both gene transcription and growth rate, implying that heterokaryotic basidiomycetes are not functionally equivalent to diploid individuals and have a higher potential for genotypic and phenotypic variation.