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Team Uses Genomic Strategies to Characterize Mountain Pine Beetle Symbiont

By Andrea Anderson

NEW YORK (GenomeWeb News) – In a study appear online last night in the Proceedings of the National Academy of Sciences, researchers from Canada and France used a combination of genomic and transcriptomic analyses to characterize Grosmannia clavigera (Gc) — a fungal symbiont of the mountain pine beetle that helps the insect infect and damage lodgepole pine trees.

In the processes, the team uncovered previously unidentified genetic patterns in the fungus, including clues about how the fungus dodges pine tree defenses.

"We developed fundamental genomic and molecular resources for functional characterization of a bark beetle-symbiotic fungus and tree pathogen," co-corresponding author Colette Breuil, a wood science researcher at the University of British Columbia, and co-authors wrote.

The mountain pine beetle has been implicated in the destruction of millions of hectares of lodgepole pine in western North America, the researchers explained. But the pest does not work alone. Past studies have shown that the mountain pine beetle depends on a specific set of microbes, including Gc, to help it infect and damage trees.

"The fungi benefit because beetles carry them through the tree bark into a new host's nutrient-rich tissues," they wrote. "The benefits to the beetle and its progeny are less clear, but the fungi make nutrients available and may detoxify host-defense metabolites."

The fungus can also kill trees on its own if enough of it is present by interfering with the movement of water from the root to the crown of the tree, Breuil told GenomeWeb Daily News. Even so, not much is known about how the fungus evades pine tree defenses on its own or in cahoots with the pine beetle.

"One surprising thing with this system is how this fungus can bypass the chemical defense system emitted by the tree," she said, "because in the tree you have a lot of chemicals which are quick toxic."

In 2009, Breuil and her colleagues reported that they had used a combination of Illumina, Roche 454, and Sanger methods to sequence a draft version of the 29.8 million base pair Gc genome.

For the current study, researchers manually finished this genome and further fleshed out the sequence data using information gleaned from RNA-sequencing analyses of seven G. clavigera strains, performed with the Illumina GAII, as well as expressed sequence tag and proteomic data.

"Using both additional pieces of information, we were able to do a better annotation of the genome," Breuil said.

Based on their data, the researchers predicted that the Gc genome houses 8,314 protein-coding genes and more than 17,200 SNPs.

Compared to fungi from 17 other taxa, the team found that the Gc seems to have expansions in several gene families, including those coding for methyltransferase and serine-peptidase enzymes. On the other hand, glycoside hydrolase, cytochrome P450, and other gene families were more compact in the pine beetle symbiont.

Even so, Breuil explained, it's still too early to know the functional consequences of these expansions and contractions.

"For the time being, it's still too early to make any hypotheses," she said. "I think we will need, really, to focus now on the functional part and do more transcriptome analyses to really understand why we have expansion of some family and contraction of other ones."

The researchers also found clues about strategies that Gc uses to sidestep tree defenses, showing that when Gc was exposed to anti-microbial compounds produced by lodgepole pine and other conifers, the fungus ramped up expression of 4,690 genes.

For instance, after 12 hours of treatment with lodgepole pine phloem extract, the team saw that the fungus had elevated expression of genes involved in sugar metabolism, oxidative stress response, and more.

Meanwhile, fungi treated with terpenoid, a compound resembling some chemicals found in the inner bark and sapwood of pine trees, bumped up their expression of genes contributing to everything from ribosome biosynthesis and messenger RNA processing to DNA repair, stability, and replication.

Because these and other gene expression changes coincide with altered histone gene expression, the researchers speculated that some of the alterations that occur in terpenoid-treated cells are a consequence of chromatin remodeling.

In the future, the team hopes to further resolve some of these pathways — for instance, by studying mutants that are missing one of the transportor genes that appears to contribute to growth in the presence of terpene.

Moreover, Breuil explained, the group plans to do additional transcriptome studies to continue teasing apart the genetic interplay between the fungus, pine trees, and pine beetles.

"By pulling all our information together, we will have a better understanding of the interaction of the three components," she said.

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