Jörg Bohlmann

University of British Columbia - Vancouver, Vancouver, British Columbia, Canada

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Publications (205)931.49 Total impact

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    ABSTRACT: The bark beetle-associated fungus Grosmannia clavigera participates in the large-scale destruction of pine forests. In the tree, it must tolerate saturating levels of toxic conifer defense chemicals (e.g. monoterpenes). The fungus can metabolize some of these compounds through the ß-oxidation pathway and use them as a source of carbon. It also uses carbon from pine triglycerides, where oleic acid is the most common fatty acid. High levels of free fatty acids, however, are toxic and can cause additional stress during host colonization. Fatty acids induce expression of neighboring genes encoding a cytochrome P450 (CYP630B18) and its redox partner, cytochrome P450 reductase (CPR2). The aim of this work was to study the function of this novel P450 system. Using LC/MS, we biochemically characterized CYP630 as a highly specific oleic acid ω-hydroxylase. We explain oleic acid specificity using protein interaction modeling. Our results underscore the importance of ω- oxidation when the main ß-oxidation pathway may be overwhelmed by other substrates such as host terpenoid compounds. Because this CYP-CPR gene cluster is evolutionarily conserved, our work has implications for metabolism studies in other fungi.
    PLoS ONE 03/2015; DOI:10.1371/journal.pone.0120119 · 3.53 Impact Factor
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    ABSTRACT: The fragrant heartwood oil of West Australian sandalwood (Santalum spicatum) contains a mixture of sesquiterpene olefins and alcohols, including variable levels of the valuable sesquiterpene alcohols, α- and β-santalol, and often high levels of E,E-farnesol. Transcriptome analysis revealed sequences for a nearly complete set of genes of the sesquiterpenoid biosynthetic pathway in this commercially valuable sandalwood species. Transcriptome sequences were produced from heartwood xylem tissue of a farnesol-rich individual tree. From the assembly of 12,537 contigs, seven different terpene synthases (TPSs), several cytochromes P450, and allylic phosphatases were identified, as well as transcripts of the mevalonic acid and methylerythritol phosphate pathways. Five of the S. spicatum TPS sequences were previously unknown. The full-length cDNA of SspiTPS4 was cloned and the enzyme functionally characterized as a multi-product sesquisabinene B synthase, which complements previous characterization of santalene and bisabolol synthases in S. spicatum. While SspiTPS4 and previously cloned sandalwood TPSs do not explain the prevalence of E,E-farnesol in S. spicatum, the genes identified in this and previous work can form a basis for future studies on natural variation of sandalwood terpenoid oil profiles. Copyright © 2014 Elsevier Ltd. All rights reserved.
    Phytochemistry 01/2015; DOI:10.1016/j.phytochem.2014.12.009 · 3.35 Impact Factor
  • Entomological Society of America Annual Meeting 2014; 11/2014
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    ABSTRACT: We developed proteome profiles for host colonizing mountain pine beetle adults, Dendroctonus ponderosae Hopkins (Coleoptera: Curculionidae). Adult insects were fed in pairs on fresh host lodgepole pine, Pinus contorta Dougl. ex Loud, phloem tissue. The proteomes of fed individuals were monitored using iTRAQ and compared to those of starved beetles, revealing 757 and 739 expressed proteins in females and males, respectively, for which quantitative information was obtained. Overall functional category distributions were similar for males and females, with the majority of proteins falling under carbohydrate metabolism (glycolysis, gluconeogenesis, citric acid cycle), structure (cuticle, muscle, cytoskeleton), and protein and amino acid metabolism. Females had 23 proteins with levels that changed significantly with feeding (p<0.05, FDR<0.20), including chaperones and enzymes required for vitellogenesis. In males, levels of 29 proteins changed significantly with feeding (p<0.05, FDR<0.20), including chaperones as well as motor proteins. Only two proteins, both chaperones, exhibited a significant change in both females and males with feeding. Proteins with differential accumulation patterns in females exhibited higher fold changes with feeding than did those in males. This difference may be due to major and rapid physiological changes occurring in females upon finding a host tree during the physiological shift from dispersal to reproduction. The significant accumulation of chaperone proteins, a cytochrome P450, and a glutathione S-transferase, indicate secondary metabolite-induced stress physiology related to chemical detoxification during early host colonization. The females' activation of vitellogenin only after encountering a host indicates deliberate partitioning of resources and a balancing of the needs of dispersal and reproduction.
    PLoS ONE 10/2014; 9(10):e110673. DOI:10.1371/journal.pone.0110673 · 3.53 Impact Factor
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    ABSTRACT: Insights from sequenced genomes of major land plant lineages have advanced research in almost every aspect of plant biology. Until recently, however, assembled genome sequences of gymnosperms have been missing from this picture. Conifers of the pine family (Pinaceae) are a group of gymnosperms that dominate large parts of the world's forests. Despite their ecological and economic importance, conifers seemed long out of reach for complete genome sequencing, due in part to their enormous genome size (20-30 Gbp) and highly repetitive nature of their genomes. Technological advances in genome sequencing and assembly enabled the recent publication of three conifer genomes: White spruce (Picea glauca), Norway spruce (P. abies), and loblolly pine (Pinus taeda). These genome sequences revealed distinctive features compared to other plant genomes, and may represent a window into the past of seed plant genomes. This update highlights recent advances, remaining challenges, and opportunities in light of the publication of the first conifer and gymnosperm genomes.
    Plant physiology 10/2014; DOI:10.1104/pp.114.248708 · 7.39 Impact Factor
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    ABSTRACT: The sesquiterpenoid rich essential oils of sandalwood (Santalum album L.) stems and roots are a highly sought commodity in the fragrance industry. Plantations of sandalwood are being established in northern Australia, however the valuable heartwood essential oils do not accumulate in substantial amounts before 10 years, while commercially viable harvests do not normally take place for at least 15 years. Inducing essential oil accumulation at an earlier stage, or increasing oil yield in mature trees, may have the potential to enhance the oil productivity of plantations. In this study, we investigated the effects of foliar application of methyl jasmonate on less than one-year-old sandalwood seedlings. Essential oil accumulation was unaffected in both stems and roots. However, at the gene transcript level, several key genes early in the biosynthesis of sandalwood oil components were induced in both leaves and stems. These results suggest that terpenoid biosynthesis in S. album does indeed respond to foliar application of methyl jasmonate, however the effects are small and the full biosynthesis of santalols is likely to be developmentally regulated. Sandalwood, Santalum album, is a small hemiparasitic tree which accumulates a mixture of sesquiterpene olefins and alcohols in the ray parenchyma of mature xylem (Jones et al. 2008). These sesquiterpenes, predominantly α-and β-santalols, constitute sandalwood oil, which is highly valued in the perfume and fragrance industry. Sandalwood trees do not normally begin to yield fragrant heartwood until approximately 10 years of age (Jones et al. 2007) suggesting that oil accumulation is developmentally controlled and heavily dependent on age. Sandalwood grows relatively slowly and even similarly aged and spaced plantation trees vary widely in the amount of heartwood oil they contain. At harvest, entire trees are uprooted as the lower part of the stems and roots contain the highest amounts of heartwood essential oils. Santalum album plantations in northern Australia are usually scheduled for harvest after 15 years, although as trees age oil contents increase (Barbour et al. 2010). Long rotation times and slow onset of heartwood development has motivated growers to seek a means to induce early sesquiterpene production in plantation sandalwood. Ideally such a process would involve a simple treatment that could be applied to leaf surfaces or through other convenient means such as irrigation in nursery facilities or plantations. Foliar application of methyl jasmonate (MeJ) has been shown to induce several different secondary metabolite biosynthetic pathways in both angiosperms and gymnosperms, with the terpenoid pathways often being highly induced (Ament et al. 2004; Miller et al. 2005; Rodriguez-Saona et al. 2001; Wang et al. 2010; Zulak et al. 2009). In the present work, initial investigation into the foliar application of MeJ was carried out on nine-month-old sandalwood saplings in the glasshouse (Figure 1A). Changes in terpenoid accumulation in roots and stems were monitored using gas chromatography-mass spectrometry (GC-MS). Concurrently, genes specific to terpene biosynthesis, along with candidate genes for pathogen defence and signalling were monitored for changes in their relative abundance by means of quantitative real-time polymerase chain reaction (qRT-PCR). By measuring these metabolic and transcript † These authors contributed equally to this work. This article can be found at
    Plant Biotechnology 10/2014; 31:585-591. DOI:10.5511/plantbiotechnology.14.1014a · 1.06 Impact Factor
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    ABSTRACT: Periodic outbreaks of spruce budworm (SBW) affect large areas of ecologically and economically important conifer forests in North America causing tree mortality and reduced forest productivity. Host resistance against SBW has been linked to growth phenology and chemical composition of foliage, but the underlying molecular mechanisms and population variation are largely unknown. Using a genomics approach, we discovered a β-glucosidase gene, Pgβglu-1, whose expression levels and function underpin natural resistance to SBW in mature white spruce (Picea glauca) trees. In phenotypically resistant trees, Pgβglu-1 transcripts were up to 1000 times more abundant compared to non-resistant trees and highly enriched in foliage. The encoded PgβGLU-1 enzyme catalyzed the cleavage of acetophenone sugar conjugates to release the aglycons piceol and pungenol. These aglycons were previously shown to be active against SBW. Levels of Pgβglu-1 transcripts and biologically active acetophenone aglycons were substantially different between resistant and non-resistant trees over time, positively correlated to each other, and highly variable in a natural white spruce population. These results suggest that expression of Pgβglu-1 and accumulation of acetophenone aglycons is a constitutive defense mechanism in white spruce. Progeny of resistant trees had higher Pgβglu-1 gene expression than non-resistant progeny indicating that the trait is heritable. With reported increases of the intensity of SBW outbreaks, influenced by climate, variation of Pgβglu-1 transcript expression, PgβGLU-1 enzyme activity, and acetophenone accumulation may serve as resistance markers to better predict impacts of SBW both in managed and wild spruce populations.This article is protected by copyright. All rights reserved.
    The Plant Journal 10/2014; 81(1). DOI:10.1111/tpj.12699 · 6.82 Impact Factor
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    ABSTRACT: The monoterpene (+)-3-carene is associated with resistance of Sitka spruce against white pine weevil, a major North American forest insect pest of pine and spruce. High and low levels of (+)-3-carene in, respectively, resistant and susceptible Sitka spruce genotypes are due to variation of (+)-3-carene synthase gene copy number, transcript and protein expression levels, enzyme product profiles, and enzyme catalytic efficiency. A family of multi-product (+)-3-carene synthase-like genes of Sitka spruce include the three (+)-3-carene synthases, PsTPS-3car1, PsTPS-3car2, PsTPS-3car3, and the (-)-sabinene synthase PsTPS-sab. Of these, PsTPS-3car2 is responsible for the relatively higher levels of (+)-3-carene in weevil-resistant trees. Here, we identified features of the PsTPS-3car1, PsTPS-3car2, PsTPS-3car3 and PsTPS-sab proteins that determine different product profiles. A series of domain swap and site-directed mutations, supported by structural comparisons, identified the amino acid in position 596 as critical for product profiles dominated by either (+)-3-carene in PsTPS-3car1, PsTPS-3car2 and PsTPS-3car3 or (-)-sabinene in PsTPS-sab. A leucine in this position promotes formation of (+)-3-carene while phenylalanine promotes (-)-sabinene. Homology modeling predicts position 596 directs product profiles through differential stabilization of the reaction intermediate. Kinetic analysis revealed position 596 also plays a role in catalytic efficiency. Mutations of position 596 with different side chain properties resulted in a series of enzymes with different product profiles, further highlighting the inherent plasticity and potential for evolution of alternative product profiles of these monoterpene synthases of conifer defence against insects.
    Journal of Biological Chemistry 07/2014; 289(34). DOI:10.1074/jbc.M114.571703 · 4.60 Impact Factor
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    ABSTRACT: Marrubium vulgare (Lamiaceae) is a medicinal plant whose major bioactive compounds, marrubiin and other labdane–related furanoid diterpenoids, have potential applications as anti–diabetics, analgesics or vasorelaxants. Metabolite and transcriptome profiling of M. vulgare leaves identified five different candidate diterpene synthases (diTPSs) of the TPS–c and TPS–e/f clades. We describe the in vitro and in vivo functional characterization of the M. vulgare diTPS family. In addition to MvEKS ent–kaurene synthase of general metabolism, we identified three diTPSs of specialized metabolism: MvCPS3 (+)-copalyl diphosphate synthase, and the functional diTPS pair MvCPS1 and MvELS. In a sequential reaction, MvCPS1 and MvELS produce a unique oxygenated diterpene scaffold 9,13-epoxy-labd-14-ene en route to marrubiin and an array of related compounds. In contrast to previously known diTPSs that introduce a hydroxyl group at carbon C–8 of the labdane backbone, the MvCPS1-catalyzed reaction proceeds via oxygenation of an intermediate carbocation at C–9, yielding the bicyclic peregrinol diphosphate. MvELS belongs to a subgroup of the diTPS TPS–e/f clade with unusual βα–domain architecture. MvELS is active in vitro and in vivo with three different prenyl diphosphate substrates forming the marrubiin precursor 9,13-epoxy-labd-14-ene, as identified by NMR, manoyl oxide and miltiradiene. MvELS fills a central position in the biosynthetic system that forms the foundation for the diverse repertoire of Marrubium diterpenoids. Co-expression of MvCPS1 and MvELS in engineered E. coli and Nicotiana benthamiana offers opportunities for producing precursors for an array of biologically active diterpenoids.This article is protected by copyright. All rights reserved.
    The Plant Journal 07/2014; DOI:10.1111/tpj.12589 · 6.82 Impact Factor
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    ABSTRACT: To successfully colonize and eventually kill pine trees, Grosmannia clavigera (Gs), the main fungal pathogen associated with the mountain-pine beetle (Dendroctonus ponderosae), has developed multiple mechanisms to overcome host tree chemical defenses, of which terpenoids are a major component. In addition to a monoterpene efflux system mediated by a recently discovered ABC transporter, Gs has genes that are highly induced by monoterpenes and that encode enzymes that modify or utilize monoterpenes (especially (+)-limonene). We showed that pine-inhabiting Ophiostomale fungi are tolerant to monoterpenes, but only a few, including Gs, are known to utilize monoterpenes as a carbon source. Gas chromatography mass spectrometry (GC/MS) revealed that Gs can modify (+)-limonene through various oxygenation pathways, producing carvone, p-mentha-2,8-dienol, perillyl alcohol and isopiperitenol. It can also degrade (+)-limonene through the C1-oxygenated pathway, producing limonene-1,2-diol as the most abundant intermediate. RNA-seq data indicated that Gs may utilize limonene 1,2-diol through beta-oxidation, then valine and TCA metabolic pathways. The data also suggested that at least two gene clusters, located in genome contigs 108 and 161, were highly induced by monoterpenes and may be involved in monoterpene degradation processes. Further, gene knock-outs indicated that limonene degradation required two distinct Baeyer-Villiger monooxygenases (BVMOs), an epoxide hydrolase and an enoyl-CoA hydratase. Our work provides information on enzyme-mediated limonene utilization or modification, and a more comprehensive understanding of the interaction between an economically important fungal pathogen and its host's defense chemicals.
    Applied and Environmental Microbiology 05/2014; 80(15). DOI:10.1128/AEM.00670-14 · 3.95 Impact Factor
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    ABSTRACT: The mountain pine beetle (MPB; Dendroctonus ponderosae Hopkins), a major pine forest pest native to western North America, has extended its range north and eastward during an ongoing outbreak. Determining how the MPB has expanded its range to breach putative barriers, whether physical (nonforested prairie and high elevation of the Rocky Mountains) or climatic (extreme continental climate where temperatures can be below -40 °C), may contribute to our general understanding of range changes as well as management of the current epidemic. Here, we use a panel of 1,536 single nucleotide polymorphisms (SNPs) to assess population genetic structure, connectivity, and signals of selection within this MPB range expansion. Biallelic SNPs in MPB from southwestern Canada revealed higher genetic differentiation and lower genetic connectivity than in the northern part of its range. A total of 208 unique SNPs were identified using different outlier detection tests, of which 32 returned annotations for products with putative functions in cholesterol synthesis, actin filament contraction, and membrane transport. We suggest that MPB has been able to spread beyond its previous range by adjusting its cellular and metabolic functions, with genome scale differentiation enabling populations to better withstand cooler climates and facilitate longer dispersal distances. Our study is the first to assess landscape-wide selective adaptation in an insect. We have shown that interrogation of genomic resources can identify shifts in genetic diversity and putative adaptive signals in this forest pest species.
    Molecular Biology and Evolution 05/2014; DOI:10.1093/molbev/msu135 · 14.31 Impact Factor
  • N. Kolosova, C. Breuil, J. Bohlmann
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    ABSTRACT: Full cDNAs encoding chitinases were cloned from lodgepole pine, the major host of the mountain pine beetle, and from interior spruce. Chitinases were induced by inoculation with bark beetle associated blue-staining fungi and insect feeding. Class I chitinases showed chitinolytic activity.
    Phytochemistry 05/2014; DOI:10.1016/j.phytochem.2014.02.006 · 3.35 Impact Factor
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    ABSTRACT: A positive relationship between genome size and intron length is observed across eukaryotes including Angiosperms plants, indicating a co-evolution of genome size and gene structure. Conifers have very large genomes and longer introns on average than most plants, but impacts of their large genome and longer introns on gene structure has not be described. Gene structure was analyzed for 35 genes of Picea glauca obtained from BAC sequencing and genome assembly, including comparisons with A. thaliana, P. trichocarpa and Z. mays. We aimed to develop an understanding of impact of long introns on the structure of individual genes. The number and length of exons was well conserved among the species compared but on average, P. glauca introns were longer and genes had four times more intronic sequence than Arabidopsis, and 2 times more than poplar and maize. However, pairwise comparisons of individual genes gave variable results and not all contrasts were statistically significant. Genes generally accumulated one or a few longer introns in species with larger genomes but the position of long introns was variable between plant lineages. In P. glauca, highly expressed genes generally had more intronic sequence than tissue preferential genes. Comparisons with the Pinus taeda BACs and genome scaffolds showed a high conservation for position of long introns and for sequence of short introns. A survey of 1836 P. glauca genes obtained by sequence capture mostly containing introns <1 Kbp showed that repeated sequences were 10x more abundant in introns than in exons. Conifers have large amounts of intronic sequence per gene for seed plants due to the presence of few long introns and repetitive element sequences are ubiquitous in their introns. Results indicate a complex landscape of intron sizes and distribution across taxa and between genes with different expression profiles.
    BMC Plant Biology 04/2014; 14(1):95. DOI:10.1186/1471-2229-14-95 · 3.94 Impact Factor
  • Philipp Zerbe, Joerg Bohlmann
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    ABSTRACT: Conifer trees, including the economically and ecologically important pine (Pinus), spruce (Picea), and fir (Abies) species, produce large amounts of oleoresin terpenoids as a defense against herbivores and pathogens. Due to the structural diversity of oleoresin terpenoids and their various chemical and physical properties, which range from solid and viscous resins to liquids and volatiles, many of these compounds are useful to humans for the production of therapeutics, fragrances and flavors, biofuels, and fine chemicals. In this chapter, we feature three examples of conifer terpenoids, the diterpene resin acids (DRA), the sesquiterpene E-α-bisabolene, and the diterpenol cis-abienol, to highlight the versatile utility of conifer terpenoids as renewable bioproducts. We focus on recent research progress on conifer terpene synthases (TPS) which produce a wealth of terpene scaffolds in nature. Our recent advances in conifer transcriptome and genome sequencing as well as metabolite analyses have accelerated discovery and definitive functional annotation of terpenoid pathway genes. New insights into the evolutionary diversification of conifer TPS, their modular organization, and dynamic expression will be fundamental to advance metabolic engineering and synthetic biology platforms for high-value terpenoids.
    Recent Advances in Phytochemistry Vol. 44, Phytochemicals – Biosynthesis, Function and Application, Edited by Reinhard Jetter, 03/2014: chapter 5: pages 85-108; Springer., ISBN: 978-3-319-04045-5
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    ABSTRACT: Studies on beetle/tree fungal symbionts typically characterize the ecological and geographic distributions of the fungal populations. There is limited understanding of the genome-wide evolutionary processes that act within and between species as such fungi adapt to different environments, leading to physiological differences and reproductive isolation. Here we assess genomic evidence for such evolutionary processes by extending our recent work on Grosmannia clavigera, which is vectored by the mountain pine beetle and jeffrey pine beetle. We report the genome sequences of an additional eleven G. clavigera sensu lato strains from the two known sibling species, Grosmannia sp. (Gs) and G. clavigera (Gc). The twelve fungal genomes are structurally similar, showing large-scale synteny within and between species. We identified 103,430 single nucleotide variations (SNVs) that separated the Grosmannia strains into divergent Gs and Gc clades, and further divided each of these clades into two subclades, one of which may represent an additional species. Comparing variable genes between these lineages, we identified truncated genes and potential pseudogenes, as well as seven genes that show evidence of positive selection. As these variable genes are involved in secondary metabolism and in detoxifying or utilizing host-tree defense chemicals (e.g. polyketide synthases, oxidoreductases, monooxygenases), their variants may reflect adaptation to the specific chemistries of the host trees Pinus contorta, P. ponderosa, and P. jeffreyi. This work provides a comprehensive resource for developing informative markers for landscape population genomics of these ecologically and economically important fungi, and an approach that could be extended to other beetle-tree associated fungi.
    Molecular Biology and Evolution 03/2014; DOI:10.1093/molbev/msu102 · 14.31 Impact Factor
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    ABSTRACT: Proanthocyanidins (PAs) are common polyphenolic polymers of plants found in foliage, fruit, bark, roots, rhizomes and seed coats that consist of flavan-3-ol units such as 2,3-trans-(+)-catechin and 2,3-cis-(-)-epicatechin. Although the biosynthesis of flavan-3-ols has been studied in angiosperms, little is known about their biosynthesis and ecological roles in gymnosperms. In this study the genes encoding leucoanthocyanidin reductase (LAR), a branch-point enzyme involved in the biosynthesis of 2,3-trans-(+)-flavan-3-ols were identified and functionally characterized in Norway spruce (Picea abies), the most widespread and economically important conifer in Europe. In addition the accumulation of flavan-3-ols and PAs was investigated in spruce saplings after wounding or inoculation with the fungal pathogen Ceratocystis polonica, which is vectored by bark beetles and is usually present during fatal beetle attacks. Monomeric and dimeric flavan-3-ols were analyzed by reverse-phase high pressure liquid chromatography, while the size and subunit composition of larger PAs were characterized using a novel acid hydrolysis method and normal phase chromatography. Only flavan-3-ol monomers with 2,3-trans stereochemistry were detected in spruce bark; dimeric and larger PAs contained flavan-3-ols with both 2,3-trans and 2,3-cis stereochemistry. Levels of monomers as well as PAs with a higher degree of polymerization increased dramatically in spruce bark after infection by C. polonica. In accordance with their role in the biosynthesis of 2,3-trans-(+)-flavan-3-ols, transcript abundance of PaLAR genes also increased significantly during fungal infection. Bioassays with C. polonica revealed that the levels of 2,3-trans-(+)-catechin and PAs which are produced in the tree in response to fungal infection inhibit C. polonica growth and can therefore be considered chemical defense compounds.
    Plant physiology 02/2014; DOI:10.1104/pp.113.232389 · 7.39 Impact Factor
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    ABSTRACT: Among the most devastating pests of Norway spruce (Picea abies) are the European spruce bark beetle (Ips typographus) and the associated pathogenic blue-stain fungus Ceratocystis polonica. Following attack and colonization, the beetle and the fungus must cope with induced host chemical defenses, such as monoterpenes that are generally thought to be toxic to both symbionts. The goal of this study was to better understand the response of Norway spruce following C. polonica inoculation at low density that does not overwhelm the tree and to identify monoterpenes mobilized toward the fungus. We inoculated healthy mature trees and monitored monoterpene profiles 2, 3, and 5 months post-inoculation. We also exposed three different C. polonica strains to the most abundant or significantly up-regulated monoterpenes to determine differences in monoterpene toxicity and resistance among strains. Total monoterpene levels, including limonene, were increased at 2 and 3 months after inoculation and had dropped after 5 months. In in vitro assays, all monoterpenes were inhibitory to C. polonica. Limonene and β-pinene were the most potent inhibitors of fungal growth. The extent of inhibition varied between the three strains tested. These results showed a defense response of Norway spruce to C. polonica, in which limonene may play a critical role in inhibiting the spread of the fungus. We also showed that differences between strains of C. polonica must be taken into account when assessing the role of the fungus in this bark beetle–symbiont system.
    European Journal of Forest Research 12/2013; 133(4). DOI:10.1007/s10342-013-0772-4 · 1.68 Impact Factor
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    ABSTRACT: The mountain pine beetle (Dendroctonus ponderosae Hopkins) is the most destructive pest of western North American pine forests. Adult males produce frontalin, an eight-carbon antiaggregation pheromone, via the mevalonate pathway, as part of several pheromones that initiate and modulate the mass attack of host trees. Frontalin acts as a pheromone, attractant, or kairomone in most Dendroctonus species, other insects, and even elephants. 6-Methylhept-6-en-2-one, a frontalin precursor, is hypothesized to originate from 10-carbon geranyl diphosphate (GPP), 15-carbon farnesyl diphosphate (FPP), or 20-carbon geranylgeranyl diphosphate (GGPP) via a dioxygenase- or cytochrome P450-mediated carbon-carbon bond cleavage. To investigate the role of isoprenyl diphosphate synthases in pheromone biosynthesis, we characterized a bifunctional GPP/FPP synthase and a GGPP synthase in the mountain pine beetle. The ratio of GPP to FPP produced by the GPP/FPP synthase was highly dependent on the ratio of the substrates isopentenyl diphosphate and dimethylallyl diphosphate used in the assay. Transcript levels in various tissues and life stages suggested that GGPP rather than GPP or FPP is used as a precursor to frontalin. Reduction of transcript levels by RNA interference of the isoprenyl diphosphate synthases identified GGPP synthase as having the largest effect on frontalin production, suggesting that frontalin is derived from a 20-carbon isoprenoid precursor rather than from the 10- or 15-carbon precursors.
    Proceedings of the National Academy of Sciences 10/2013; 110(47). DOI:10.1073/pnas.1316498110 · 9.81 Impact Factor
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    ABSTRACT: The mountain pine beetle (MPB, Dendroctonus ponderosae Hopkins) is a significant pest of western North American pine forests. This beetle responds to pheromones and host volatiles in order to mass attack and thus overcome the terpenoid chemical defences of its host. The ability of MPB antennae to rapidly process odorants is necessary to avoid odorant receptor saturation and thus the enzymes responsible for odorant clearance are an important aspect of host colonization. An antenna-specific cytochrome P450, DponCYP345E2, is the most highly expressed transcript in adult MPB antenna. In in vitro assays with recombinant enzyme, DponCYP345E2 used several pine host monoterpenes as substrates, including (+)-(3)-carene, (+)-β-pinene, (-)-β-pinene, (+)-limonene, (-)-limonene, (-)-camphene, (+)-α-pinene, (-)-α-pinene, and terpinolene. The substrates were epoxidized or hydroxylated, depending upon the substrate. To complement DponCYP345E2, we also functionally characterized the NADPH-dependent cytochrome P450 reductase and the cytochrome b5 from MPB. DponCYP345E2 is the first cytochrome P450 to be functionally characterized in insect olfaction and in MPB.
    Insect biochemistry and molecular biology 10/2013; 43(12). DOI:10.1016/j.ibmb.2013.10.001 · 3.25 Impact Factor

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  • 2000–2015
    • University of British Columbia - Vancouver
      • • Michael Smith Laboratories
      • • Department of Wood Science
      • • Department of Forest Sciences
      Vancouver, British Columbia, Canada
  • 2013
    • University of Northern British Columbia
      Prince George, British Columbia, Canada
  • 2008–2011
    • Government of British Columbia, Canada
      Vancouver, British Columbia, Canada
  • 2000–2011
    • Max Planck Institute for Chemical Ecology
      • Department of Biochemistry
      Jena, Thuringia, Germany
  • 2005–2009
    • Upsher-Smith Laboratories, Ing.
      Minneapolis, Minnesota, United States
  • 2006
    • Simon Fraser University
      • Department of Biological Sciences
      Burnaby, British Columbia, Canada
  • 2003
    • Purdue University
      • Department of Horticulture and Landscape Architecture
      West Lafayette, IN, United States
  • 1996–1999
    • Washington State University
      • Institute of Biological Chemistry
      Pullman, WA, United States
  • 1995
    • Technische Universität Braunschweig
      • Institute of Genetics
      Brunswyck, Lower Saxony, Germany