Dr. J. Bohlmann

RESEARCH PROGRAM

1. Summary: Secondary metabolism; Plant defense against insects and insect-associated pathogens; Forestry and grapevine genomics

Research in my group is developing a genomic and functional biochemical characterization of conifer defense systems that are active against insect pests and insect-associated fungal pathogens. Some of this research is based on our studies of the molecular biochemistry of plant terpenoid secondary metabolism (see below). Our research has laid the foundation for Canada’s first large-scale forestry genomics program (Treenomix, www.treenomix.ca; funded by Genome Canada, Genome British Columbia and the Province of British Columbia) of which I have been a project leader since 2001. With the Treenomix project I have extended my program to include research on poplar as a model system for tree biology. My lab has been leading genomics research on defense against insects in long-lived, woody plants. In 2006, our forestry genomics program has been refunded with a new project “Conifer Forest Health” genomics. While my research program is primarily designed as fundamental and curiosity-driven research, the results of our research program can potentially be translated into enormous economic, social, and environmental benefit through the long-term sustainability of Canada’s forests. These forests are already under pressure from insect pests, of which the current mountain pine beetle epidemic in British Columbia is just one example. Under predicted scenarios of global climate change forest insect pests will continue to increase as a major threat to Canada’s forests. Our research program on forest health involves collaboration with entomologists targeting the molecular biochemistry of bark beetle pheromone systems (initially supported with an HFSP young investigators award), and collaboration with microbiologists targeting the interacting genomes of bark beetle-associated fungal pathogens and pine host trees (supported with an NSERC strategic grant). The original foundation of my research in plant secondary metabolism, specifically my research on the molecular biochemistry of terpenoid natural products, allowed me to develop several other successful lines of related research. These research activities include research on grapevine (Vitis vinifera) flavor and aroma biochemistry (supported with a Genome Canada and Genome Spain international program grant), collaborative research on floral scent biochemistry, and research on the role and regulation of terpenoids in Arabidopsis thaliana.

2. Research on the molecular genetic, biochemical, and chemical characterization of terpenoid defenses in conifers and poplars against insect pests and pathogens; Terpenoid synthases and cytochrome P450 enzymes

Insect pests and pathogens cause annual losses of billions of dollars to conifer-based forest economies in North America and Europe. To address this issue, my group has established a program on the molecular genetic and biochemical characterization of chemical defenses in conifers. As some of the oldest and longest living plants, conifers have evolved complex, terpenoid-based defense mechanisms against insect herbivores and pathogens. These chemical defenses occur both as preformed defenses and as inducible defenses. In my postdoctoral research I characterized a family of terpene synthase (TPS) genes and enzymes from the conifer grand fir, Abies grandis. The TPS are responsible for the formation of oleoresin terpenes in this system. I isolated cDNAs for a multiple member TPS gene family, expressed the cDNAs in E. coli for in vitro characterization of recombinant enzymes, analyzed TPS gene expression in wound-induced trees and reconstructed a phylogeny of the plant TPS gene family. This work provided an understanding of highly complex defense mechanisms in conifers, relevant to forest biotechnology, and can lead to new strategies for pest control. With my group at at UBC, I have build on our knowledge from the grand fir system to explore more comprehensively terpenoid defenses in the economically important conifer species of spruce (Picea spp.), pines (Pinus spp.), and Douglas fir (Pseudotsuga menziesii). Like other conifers, species of spruce accumulate terpenoids constitutively in preformed cortical resin ducts. In addition, upon insect feeding or pathogen challenge, these trees also respond with an unusual induced de novo differentiation of traumatic resin ducts (TDs) in the developing xylem. Research in my group showed that treatment of trees with the defense signal compound methyl jasmonate (MeJA) induces increased resin terpenoid biosynthesis and traumatic resin accumulation, as well as de novo development of TDs. A suite of chemically defined elicitors has been evaluated for their effect on activation of spruce defenses in pretreatment studies. To further analyze in detail the molecular regulation of resin terpenoid defenses in species of spruce, we isolated cDNAs for a large family of TPS genes from Norway spruce (P. abies) and Sitka spruce (P. sitchensis). We functionally characterized many of these genes and analyzed gene expression in response to insect herbivory, wounding, or treatment of trees with MeJA. We then found that Norway spruce and Sitka spruce trees also respond to treatment with MeJA and to insect attack with the emission of volatile organic compounds. The emission of volatiles was in addition to activation of direct, resin-based terpenoid defenses. Such volatiles can serve as info-chemicals in indirect defense systems of plants. Based on a detailed understanding of the signals and mechanisms of induced direct and indirect terpenoid defenses in spruce we are now developing new strategies for protection of forest trees against insect pests. To complement our research on conifer-insect interactions with a comparative analysis of a conifer-pathogen system, we have developed in parallel a successful characterization of chemical defenses of Douglas fir (Pseudotsuga menziesii) trees that are hosts to certain root rot fungi. Recently we have expanded our program on terpenoid biochemistry in conifers with the discovery of a new group of cytochrome P450 genes in spruce and in loblolly pine (Pinus taeda). Using an integrated functional genomics, FL-cDNA, and metabolite profiling approach, we successfully identified and characterized a new gymnosperm cytochrome P450 gene. We characterized the corresponding recombinant P450 enzyme as an unusual multifunctional and multi-substrate diterpene oxidase with a metabolic function in diterpene resin acid biosynthesis. Other research with conifers that is ongoing in my group has been studying molecular and biochemical mechanisms of induced defenses in lodgepole pine (Pinus contorta) against its fungal pathogen (Ophiostoma clavigerum), a blue stain fungus that is associated with the mountain pine beetle (Dendroctonus ponderosa). The mountain pine beetle epidemic is the largest recorded bark beetle outbreak in Canada causing major economic losses to the forest industry. In a comparative analysis of terpenoid defenses in gymnosperm and angiosperm forest trees, we have investigated indirect tritrophic defense systems in the poplar (Populus) / forest tent caterpillar (FTC) system. In poplars, we identified spatial and temporal patterns of FTC-induced systemic volatile emissions and the TPS gene and enzyme responsible for FTC-induced, diurnal emission of ()-germacrene D, a sesquiterpene.


3. Forestry Genomics: Treenomix, Canada’s first large-scale forestry genomics project (2001 – 2005); Conifer Forest Health Genomics (since 2006) funded by Genome Canada, Genome British Columbia, and the Province of British Columbia

Our studies on chemical defense against insects and pathogens in conifers and poplars provided a biological and functional foundation for Canada’s first large-scale Forestry Genome project “Treenomix”(www.treenomix.ca) and a new genomics project, “Conifer Forest Health”. I am directing the “Conifer Forest Health” project together with Dr. Kermit Ritland (UBC, Dept. Forest Sciences). Collaborators at UBC are Drs. Sally. Aitken, Gary Bull, Thomas Maness, Shawn Mansfield, Paul Wood. Our international partners are at the University of California, Davis (Dr. David Neale), at the Max Planck Institute for Chemical Ecology, Germany (Drs. Jonathan Gershenzon and Michael Phillips), at the Umeå Plant Sciences Centre, Sweden (Drs. Rhishi Bhalerao, Thomas Moritz, Ove Nilsson, Göran Sandberg, Björn Sundberg, and others), and Forest Research Institute in the UK (Dr. Steven Lee). In our forestry genomics program we are using a combination of genomic, proteomic, and metabolite profiling tools to decipher constitutive and induced defense mechanisms and genetic resistance in forest trees, with a primary emphasis on species of spruce, Sitka spruce (Picea sitchensis), White spruce (P. glauca), and Norway spruce (P. abies). In addition, we are using Arabidopsis thaliana and poplar as reference and model systems. Our program has developed essential genomic tools to increase the scientific community’s understanding about trees’ built-in defense mechanisms against insect pests, insect-assocaited pathogens, and environmental stress. The program uses a vertically integrated approach that transfers information between the diverse species of spruce, species of pines, poplar, and Arabidopsis. Specifically, cDNA ESTs and full-length cDNAs from spruce and poplar have been sequenced, obtaining a database and cDNA clone collection of more than 350,000 ESTs from each tree species. The project builds further upon the whole-genome sequence of Arabidopsis and poplar, and on the ability of these plants to display insect-induced defense responses. EST sequences are used to study gene expression underlying important traits in tree defense using microarrays, identifying similarities and differences across resistant and susceptible genotypes. To this end we have developed a 15.5k poplar cDNA microarray and a 22k spruce cDNA microarray. We have identified differentially expressed proteins related to defense against insects in spruce, poplar and Arabidopsis by 2-D gel analysis coupled with Q-TOF and MALDI-TOF MS, and by ICAT and iTRAQ technologies. For spruce and poplar libraries of more than ten thousand genes have been cloned as full-length (FL) cDNAs and in each species more than 4,000 non-redundant FL-cDNAs have been completely sequenced for future functional characterization. Bioinformatic comparisons are being made of sequence changes across this group of species, and correlated with trait evolution. The project is also identifying genetic markers to help in seed orchard and breeding programs, and to locate and identity major genes underlying economically important traits in forestry. As a contribution to the whole-genome sequencing of poplar, a physical map of this tree has been generated from the fingerprinting of 48,000 BAC clones and 96,000 BAC end-sequences. With this work and the FL-cDNAs from poplar, our project has contributed to an international effort with the US Department of Energy (DOE) Joint Genomics Institute (JGI), the Umeå Plant Science Center in Sweden, and several other international partners to sequence, assemble and annotate the poplar genome. A first draft poplar genome sequence was released in September of 2004. This work in tree genomics has been the first of its kind in Canada and is internationally recognized to be at the forefront of forestry genomics. Most of our research on functional genomics is now targeted at identifying and characterizing forest tree defense systems against insects and insect-associated fungi. Specifically we are identifying defense and possible resistance mechanisms of Sitka spruce and white spruce against the white pine weevil and spruce budworm; defense mechanisms of lodgepole pine against the bark beetle associated blue stain fungus Ophiostoma clavigerum, and defense mechanisms of poplars against forest tent caterpillar and willow borer.

4. Bark beetle pheromone de novo biosynthesis

Bark beetles use a complex pheromone communication system in the successful attack of conifer host trees. Myrcene is an acyclic monoterpene that has long been associated with pheromone synthesis in pine bark beetles (e.g. the pine engraver, Ips pini). The logical source of myrcene for bark beetle pheromone biosynthesis has been the oleoresin of the host pines. Monoterpene synthases, which convert geranyl diphosphate to monoterpenoid natural products, have been frequently characterized from conifers and other plants. However, they have not previously been found in animals. We discovered a monoterpene synthase activity in two populations of an insect, I. pini. Cell-free assays of I. pini revealed that 3H-geranyl diphosphate is converted to 3H-myrcene in whole-body extracts from male, but not female, I. pini. Furthermore, the enzyme activity in males can be induced by prior treatment with juvenile hormone III (JH III) or by feeding on phloem from the host trees, Jeffrey pine (Pinus jeffreyi) or red pine (Pinus resinosa). The sex-specificity and endocrine induction of this activity argue for its involvement in the biosynthesis of monoterpenoid pheromones mediated by enzymes from I. pini rather than from microbial symbionts. This work has been in collaboration with Dr. Steven Seybold of the USDA Forests Service, Davis. We have received funding from the international HFSP young investigators program to support this interdisciplinary collaboration between our terpenoid biochemistry group and Dr. Seybold’s entomology group.

5. Molecular biochemistry and genomics of floral scents and fruit flavor and aroma

In collaboration with Dr. Natalia Dudareva (Purdue University, USA), my lab has contributed to the discovery of floral scent genes in snapdragon (Antirrhinum majus) and identified biochemical functions of the encoded enzymes and biochemical and molecular mechanisms of terpenoid floral scent emission in this system. As part of my faculty affiliation with the UBC Wine Research Centre, I started in 2002 a project on the biochemical characterization of monoterpenoid and sesquiterpenoid aroma genes in grape vine (Vitis vinifera). Relevant candidate genes have been identified in EST databases. Selected genes have been functionally identified [()--terpineol synthase; (+) valencene synthase; ()-germacrene D synthase]. Our research with grape has recently received major support with a 3.1 million dollar international grant from Genome Canada / Genome Spain. This new 3-year project started in the spring of 2004, under the direction of Dr. Steven Lund (UBC), and in collaboration with Dr. Patricia Bowen (PARC, Penticton) with international partners in Spain.


6. Functional genomics of terpenoid in Arabidopsis thaliana

While research on conifers and forest health is the major activity of my program, my group is also studying a few selected aspects of low molecular weight terpenoids in Arabidopsis thaliana. Based on a genomics approach, we started identification and functional characterization of TPS genes involved in secondary metabolism in Arabidopsis. In a systematic analysis of the Arabidopsis genome sequence we discovered a surprisingly large family of terpene synthase (AtTPS) genes in this model plant system. The genome-wide analysis of the AtTPS gene family provided novel insights into genome organization and evolution of the plant TPS gene family. The AtTPS genes resembled TPS genes previously described for plants that produce an array of monoterpene, sesquiterpene, and diterpene secondary metabolites in a context of chemical interaction with other organisms. While terpenoid secondary metabolites of this type had not been described for Arabidopsis prior to our work, our discovery of a large and highly conserved AtTPS gene family suggested functional roles of TPS genes and terpenoid secondary metabolites in Arabidopsis. This surprising finding may be implicative of common roles for low molecular weight terpenoid compounds in plants. Recent gene-specific microarray analysis of the complete AtTPS gene family revealed differential and often organ-specific expression of AtTPS genes in Arabidopsis and is guiding ongoing functional characterization of AtTPS genes and terpenoid natural products in this system.

7. Collaborations, Funding, Facilities

Our research projects involve national and international collaborations, several of which are mentioned in the summary of research (see above).

Funding for our research is from the Natural Sciences and Engineering Research Council of Canada (NSERC) through its Discovery and Strategic programs and student and postdoctoral fellowship programs, the Canadian Foundation for Innovation (CFI), the British Columbia Knowledge and Development Funds (BCKDF), Genome Canada, Genome BC, the Province of British Columbia, and other sources.

My group uses laboratory space in the new Michael Smith Laboratories, in the adjacent National Centre of Excellence (NCE), and in the Forest Sciences Centre. We have modern equipment and facilities for genomics, microarray gene expression profiling, proteomics sample preparation, plant molecular biology, enzyme biochemistry, and analytics. Together with Dr. Hennie van Vuuren, UBC Wine Research Centre, I have developed a dedicated mass spectrometry laboratory for analysis of small molecules (e.g. plant secondary metabolites, signaling compounds) equipped with new GC/MS, GC/FID, radio-GC, and LC/MS instruments.

Keeling CI, Dullat HK, Yuen M, Ralph SG, Jancsik S, Bohlmann J. Identification and functional characterization of monofunctional ent-copalyl diphosphate and ent-kaurene synthases in white spruce (Picea glauca) reveal different patterns for diterpene synthase evolution for primary and secondary metabolism in gymnosperms. Plant Physiol . (2010).

Zulak KG, Bohlmann J Terpenoid biosynthesis and specialized vascular cells of conifer defense Journal of Integrative Plant Biology (In Press) . (2010).

Zulak KG, Lippert DN, Kuzyk MA, Domanski D, Chou T, Borchers CH, Bohlmann J. Targeted proteomics using selected reaction monitoring reveals the induction of specific terpene synthases in a multi-level study of methyl jasmonate-treated Norway spruce (Picea abies). Plant J 60(6):1015-30. (2009).

Philippe RN, Ralph SG, Kulheim C, Jancsik SI, Bohlmann J. Poplar defense against insects: genome analysis, full-length cDNA cloning, and transcriptome and protein analysis of the poplar Kunitz-type protease inhibitor family. New Phytol 184(4):865-84. (2009).

Diguistini S, Liao NY, Platt D, Robertson G, Seidel M, Chan SK, Docking TR, Birol I, Holt RA, Hirst M, Mardis E, Marra MA, Hamelin RC, Bohlmann J, Breuil C, Jones SJ. De novo genome sequence assembly of a filamentous fungus using Sanger, 454 and Illumina sequence data. Genome Biol 10(9):R94. (2009).

Hamberger B, Hall D, Yuen M, Oddy C, Hamberger B, Keeling CI, Ritland C, Ritland K, Bohlmann J. Targeted isolation, sequence assembly and characterization of two white spruce (Picea glauca) BAC clones for terpenoid synthase and cytochrome P450 genes involved in conifer defence reveal insights into a conifer genome. BMC Plant Biol 9106. (2009).

Bohlmann J, Gershenzon J. Old substrates for new enzymes of terpenoid biosynthesis. Proc Natl Acad Sci U S A 106(26):10402-3. (2009).

Dafoe NJ, Zamani A, Ekramoddoullah AK, Lippert D, Bohlmann J, Constabel CP. Analysis of the poplar phloem proteome and its response to leaf wounding. J Proteome Res 8(5):2341-50. (2009).

Martin DM, Toub O, Chiang A, Lo BC, Ohse S, Lund ST, Bohlmann J. The bouquet of grapevine (Vitis vinifera L. cv. Cabernet Sauvignon) flowers arises from the biosynthesis of sesquiterpene volatiles in pollen grains. Proc Natl Acad Sci U S A 106(17):7245-50. (2009).

Lippert DN, Ralph SG, Phillips M, White R, Smith D, Hardie D, Gershenzon J, Ritland K, Borchers CH, Bohlmann J. Quantitative iTRAQ proteome and comparative transcriptome analysis of elicitor-induced Norway spruce (Picea abies) cells reveals elements of calcium signaling in the early conifer defense response. Proteomics 9(2):350-67. (2009).

Tsui CKM, Feau N, Ritland CE, Massoumi Alamouti S, DiGuistini S, Bohlmann J, Breuil C, and RC Hamelin Characterization of microsatellite loci in the fungus Grosmannia clavigera, a pine pathogen associated with the mountain pine beetle. Molecular Ecology Resources 91500-1503. (2009).

Lippert D, Yuen M, and J Bohlmann Spruce Proteome DB: A resource for conifer proteomics research. Tree Genetics & Genomes (5):723 – 727. (2009).

Liewlaksaneeyanawin C, Zhuang J, Tang M, Farzaneh N, Lueng G, Oddy C, Findlay S, Ritland CE, Bohlmann J, and K Ritland Identification and validation of COS markers in Pinaceae Tree Genetics & Genomes (5):247-255. (2009).

Ralph SG, Chun HJ, Kolosova N, Cooper D, Oddy C, Ritland CE, Kirkpatrick R, Moore R, Barber S, Holt RA, Jones SJ, Marra MA, Douglas CJ, Ritland K, Bohlmann J. A conifer genomics resource of 200,000 spruce (Picea spp.) ESTs and 6,464 high-quality, sequence-finished full-length cDNAs for Sitka spruce (Picea sitchensis). BMC Genomics 9484. (2008).

Jones CG, Keeling CI, Ghisalberti EL, Barbour EL, Plummer JA, Bohlmann J. Isolation of cDNAs and functional characterisation of two multi-product terpene synthase enzymes from sandalwood, Santalum album L. Arch Biochem Biophys 477(1):121-30. (2008).

Godard KA, White R, Bohlmann J. Monoterpene-induced molecular responses in Arabidopsis thaliana. Phytochemistry 69(9):1838-49. (2008).

Bohlmann J, Keeling CI. Terpenoid biomaterials. Plant J 54(4):656-69. (2008).

Ehlting J, Chowrira SG, Mattheus N, Aeschliman DS, Arimura G, Bohlmann J. Comparative transcriptome analysis of Arabidopsis thaliana infested by diamond back moth (Plutella xylostella) larvae reveals signatures of stress response, secondary metabolism, and signalling. BMC Genomics 9154. (2008).

Ralph SG, Chun HJ, Cooper D, Kirkpatrick R, Kolosova N, Gunter L, Tuskan GA, Douglas CJ, Holt RA, Jones SJ, Marra MA, Bohlmann J. Analysis of 4,664 high-quality sequence-finished poplar full-length cDNA clones and their utility for the discovery of genes responding to insect feeding. BMC Genomics 957. (2008).

Keeling CI, Weisshaar S, Lin RP, Bohlmann J. Functional plasticity of paralogous diterpene synthases involved in conifer defense. Proc Natl Acad Sci U S A 105(3):1085-90. (2008).

Holliday JA, Ralph SG, White R, Bohlmann J, Aitken SN. Global monitoring of autumn gene expression within and among phenotypically divergent populations of Sitka spruce (Picea sitchensis). New Phytol 178(1):103-22. (2008).

Bohlmann J Insect-induced terpenoid defenses in spruce. ln: "Induced Plant Resistance to Herbivory" A. Schaller (ed.) Springer Science 137-187. (2008).

Keeling CI and Bohlmann J Terpenoids in plants Wiley Encyclopedia of Chemical Biology, doi: 10.1002/9780470048672.webcb596 . (2008).

Souza Cde A, Barbazuk B, Ralph SG, Bohlmann J, Hamberger B, Douglas CJ. Genome-wide analysis of a land plant-specific acyl:coenzyme A synthetase (ACS) gene family in Arabidopsis, poplar, rice and Physcomitrella. New Phytol 179(4):987-1003. (2008).

Godard KA, Byun-McKay A, Levasseur C, Plant A, Seguin A, Bohlmann J. Testing of a heterologous, wound- and insect-inducible promoter for functional genomics studies in conifer defense. Plant Cell Rep 26(12):2083-90. (2007).

Peng FY, Reid KE, Liao N, Schlosser J, Lijavetzky D, Holt R, Marti­nez Zapater JM, Jones S, Marra M, Bohlmann J, Lund ST. Generation of ESTs in Vitis vinifera wine grape (Cabernet Sauvignon) and table grape (Muscat Hamburg) and discovery of new candidate genes with potential roles in berry development. Gene 402(1):40-50. (2007).

Phillips MA, Walter MH, Ralph SG, Dabrowska P, Luck K, Urós EM, Boland W, Strack D, Rodríguez-Concepción M, Bohlmann J, Gershenzon J. Functional identification and differential expression of 1-deoxy-D-xylulose 5-phosphate synthase in induced terpenoid resin formation of Norway spruce (Picea abies). Plant Mol Biol 65(3):243-57. (2007).

Behnke K, Ehlting B, Teuber M, Bauerfeind M, Louis S, Hänsch R, Polle A, Bohlmann J, Schnitzler JP. Transgenic, non-isoprene emitting poplars don't like it hot. Plant J 51(3):485-99. (2007).

Miranda M, Ralph SG, Mellway R, White R, Heath MC, Bohlmann J, Constabel CP. The transcriptional response of hybrid poplar (Populus trichocarpa x P. deltoides) to infection by Melampsora medusae leaf rust involves induction of flavonoid pathway genes leading to the accumulation of proanthocyanidins. Mol Plant Microbe Interact 20(7):816-31. (2007).

Ralph SG, Jancsik S, Bohlmann J. Dirigent proteins in conifer defense II: Extended gene discovery, phylogeny, and constitutive and stress-induced gene expression in spruce (Picea spp.). Phytochemistry 68(14):1975-91. (2007).

Kelleher CT, Chiu R, Shin H, Bosdet IE, Krzywinski MI, Fjell CD, Wilkin J, Yin T, DiFazio SP, Ali J, Asano JK, Chan S, Cloutier A, Girn N, Leach S, Lee D, Mathewson CA, Olson T, O'connor K, Prabhu AL, Smailus DE, Stott JM, Tsai M, Wye NH, Yang GS, Zhuang J, Holt RA, Putnam NH, Vrebalov J, Giovannoni JJ, Grimwood J, Schmutz J, Rokhsar D, Jones SJ, Marra MA, Tuskan GA, Bohlmann J, Ellis BE, Ritland K, Douglas CJ, Schein JE. A physical map of the highly heterozygous Populus genome: integration with the genome sequence and genetic map and analysis of haplotype variation. Plant J 50(6):1063-78. (2007).

Huber DP, Erickson ML, Leutenegger CM, Bohlmann J, Seybold SJ. Isolation and extreme sex-specific expression of cytochrome P450 genes in the bark beetle, Ips paraconfusus, following feeding on the phloem of host ponderosa pine, Pinus ponderosa. Insect Mol Biol 16(3):335-49. (2007).

DiGuistini S, Ralph SG, Lim YW, Holt R, Jones S, Bohlmann J, Breuil C. Generation and annotation of lodgepole pine and oleoresin-induced expressed sequences from the blue-stain fungus Ophiostoma clavigerum, a Mountain Pine Beetle-associated pathogen. FEMS Microbiol Lett 267(2):151-8. (2007).

Friedmann M, Ralph SG, Aeschliman D, Zhuang J, Ritland K, Ellis BE, Bohlmann J, Douglas CJ. Microarray gene expression profiling of developmental transitions in Sitka spruce (Picea sitchensis) apical shoots. J Exp Bot 58(3):593-614. (2007).

Lippert D, Chowrira S, Ralph SG, Zhuang J, Aeschliman D, Ritland C, Ritland K, Bohlmann J. Conifer defense against insects: proteome analysis of Sitka spruce (Picea sitchensis) bark induced by mechanical wounding or feeding by white pine weevils (Pissodes strobi). Proteomics 7(2):248-70. (2007).

Ralph SG, Hudgins JW, Jancsik S, Franceschi VR, Bohlmann J. Aminocyclopropane carboxylic acid synthase is a regulated step in ethylene-dependent induced conifer defense. Full-length cDNA cloning of a multigene family, differential constitutive, and wound- and insect-induced expression, and cellular and subcellular localization in spruce and Douglas fir. Plant Physiol 143(1):410-24. (2007).

Berube Y, Zhuang J, Ralph S, Rungis D, Bohlmann J, and K Ritland Characterization of EST-SSRs in loblolly pine and spruce. Tree Genetics & Genomics 3: 251-259 . (2007).

Philippe R.N. and Bohlmann J Poplar defense against insect herbivores. Canadian Journal of Botany 85: 111-1126 . (2007).

Hamberger B, Bohlmann J. Cytochrome P450 mono-oxygenases in conifer genomes: discovery of members of the terpenoid oxygenase superfamily in spruce and pine. Biochem Soc Trans 341209-14. (2006).

Keeling CI, Bohlmann J. Diterpene resin acids in conifers. Phytochemistry 67(22):2415-23. (2006).

Tuskan GA, Difazio S, Jansson S, Bohlmann J, Grigoriev I, Hellsten U, Putnam N, Ralph S, Rombauts S, Salamov A, Schein J, Sterck L, Aerts A, Bhalerao RR, Bhalerao RP, Blaudez D, Boerjan W, Brun A, Brunner A, Busov V, Campbell M, Carlson J, Chalot M, Chapman J, Chen GL, Cooper D, Coutinho PM, Couturier J, Covert S, Cronk Q, Cunningham R, Davis J, Degroeve S, Déjardin A, Depamphilis C, Detter J, Dirks B, Dubchak I, Duplessis S, Ehlting J, Ellis B, Gendler K, Goodstein D, Gribskov M, Grimwood J, Groover A, Gunter L, Hamberger B, Heinze B, Helariutta Y, Henrissat B, Holligan D, Holt R, Huang W, Islam-Faridi N, Jones S, Jones-Rhoades M, Jorgensen R, Joshi C, Kangasjärvi J, Karlsson J, Kelleher C, Kirkpatrick R, Kirst M, Kohler A, Kalluri U, Larimer F, Leebens-Mack J, Leplé JC, Locascio P, Lou Y, Lucas S, Martin F, Montanini B, Napoli C, Nelson DR, Nelson C, Nieminen K, Nilsson O, Pereda V, Peter G, Philippe R, Pilate G, Poliakov A, Razumovskaya J, Richardson P, Rinaldi C, Ritland K, Rouzé P, Ryaboy D, Schmutz J, Schrader J, Segerman B, Shin H, Siddiqui A, Sterky F, Terry A, Tsai CJ, Uberbacher E, Unneberg P, Vahala J, Wall K, Wessler S, Yang G, Yin T, Douglas C, Marra M, Sandberg G, Van de Peer Y, Rokhsar D. The genome of black cottonwood, Populus trichocarpa (Torr. & Gray). Science 313(5793):1596-604. (2006).

Hudgins JW, Ralph SG, Franceschi VR, and J Bohlmann Ethylene in induced conifer defense response: FLcDNA cloning, wound-induced protein expression, and cellular and subcellular localization of 1-aminocyclopropane-carboxylic acid oxidase (ACO) in specialized resin duct cells and polyphenolic phloem parechnyma cells. Planta (224):865-877. (2006).

Seybold SJ, Huber DPW, Lee LC, and J Bohlmann Pine monoterpenes and pine bark beetles: A marriage of convenience for defense and chemical communication. Phytochemistry Reviews (5):143-178. (2006).

Phillips M, Bohlmann J, and J Gershenzon Molecular regulation of induced terpenoid biosynthesis in conifers. Phytochemistry Reviews (5):179-189. (2006).

Ralph SG, Yueh H, Friedmann M, Aeschliman D, Zeznik JA, Nelson CC, Butterfield YS, Kirkpatrick R, Liu J, Jones SJ, Marra MA, Douglas CJ, Ritland K, Bohlmann J. Conifer defence against insects: microarray gene expression profiling of Sitka spruce (Picea sitchensis) induced by mechanical wounding or feeding by spruce budworms (Choristoneura occidentalis) or white pine weevils (Pissodes strobi) reveals large-scale changes of the host transcriptome. Plant Cell Environ 29(8):1545-70. (2006).

Ro DK, Bohlmann J. Diterpene resin acid biosynthesis in loblolly pine (Pinus taeda): Functional characterization of abietadiene/levopimaradiene synthase (PtTPS-LAS) cDNA and subcellular targeting of PtTPS-LAS and abietadienol/abietadienal oxidase (PtAO, CYP720B1). Phytochemistry 67(15):1572-8. (2006).

Ro DK, Ehlting J, Keeling CI, Lin R, Mattheus N, Bohlmann J. Microarray expression profiling and functional characterization of AtTPS genes: duplicated Arabidopsis thaliana sesquiterpene synthase genes At4g13280 and At4g13300 encode root-specific and wound-inducible (Z)-gamma-bisabolene synthases. Arch Biochem Biophys 448(1):104-16. (2006).

Ralph S, Oddy C, Cooper D, Yueh H, Jancsik S, Kolosova N, Philippe RN, Aeschliman D, White R, Huber D, Ritland CE, Benoit F, Rigby T, Nantel A, Butterfield YS, Kirkpatrick R, Chun E, Liu J, Palmquist D, Wynhoven B, Stott J, Yang G, Barber S, Holt RA, Siddiqui A, Jones SJ, Marra MA, Ellis BE, Douglas CJ, Ritland K, Bohlmann J. Genomics of hybrid poplar (Populus trichocarpax deltoides) interacting with forest tent caterpillars (Malacosoma disstria): normalized and full-length cDNA libraries, expressed sequence tags, and a cDNA microarray for the study of insect-induced defences in poplar. Mol Ecol 15(5):1275-97. (2006).

Gilchrist EJ, Haughn GW, Ying CC, Otto SP, Zhuang J, Cheung D, Hamberger B, Aboutorabi F, Kalynyak T, Johnson L, Bohlmann J, Ellis BE, Douglas CJ, Cronk QC. Use of Ecotilling as an efficient SNP discovery tool to survey genetic variation in wild populations of Populus trichocarpa. Mol Ecol 15(5):1367-78. (2006).

Byun-McKay A, Godard KA, Toudefallah M, Martin DM, Alfaro R, King J, Bohlmann J, Plant AL. Wound-induced terpene synthase gene expression in Sitka spruce that exhibit resistance or susceptibility to attack by the white pine weevil. Plant Physiol 140(3):1009-21. (2006).

Lund ST, Bohlmann J. The molecular basis for wine grape quality--a volatile subject. Science 311(5762):804-5. (2006).

Ralph S, Park JY, Bohlmann J, Mansfield SD. Dirigent proteins in conifer defense: gene discovery, phylogeny, and differential wound- and insect-induced expression of a family of DIR and DIR-like genes in spruce (Picea spp.). Plant Mol Biol 60(1):21-40. (2006).

Keeling CI, Bohlmann J. Genes, enzymes and chemicals of terpenoid diversity in the constitutive and induced defence of conifers against insects and pathogens. New Phytol 170(4):657-75. (2006).

Ritland K, Ralph S, Lippert D, Rungis D, and J Bohlmann A new direction in conifer genomics. In: “Landscapes, Genomics and Transgenic Conifer Forests” Claire Williams (ed.) Springer Publishers, Dordrecht Netherlands 75-84. (2006).

Huber DP, Philippe RN, Madilao LL, Sturrock RN, Bohlmann J. Changes in anatomy and terpene chemistry in roots of Douglas-fir seedlings following treatment with methyl jasmonate. Tree Physiol 25(8):1075-83. (2005).

Ehlting J, Mattheus N, Aeschliman DS, Li E, Hamberger B, Cullis IF, Zhuang J, Kaneda M, Mansfield SD, Samuels L, Ritland K, Ellis BE, Bohlmann J, Douglas CJ. Global transcript profiling of primary stems from Arabidopsis thaliana identifies candidate genes for missing links in lignin biosynthesis and transcriptional regulators of fiber differentiation. Plant J 42(5):618-40. (2005).

Huber DP, Philippe RN, Godard KA, Sturrock RN, Bohlmann J. Characterization of four terpene synthase cDNAs from methyl jasmonate-induced Douglas-fir, Pseudotsuga menziesii. Phytochemistry 66(12):1427-39. (2005).

Ro DK, Arimura G, Lau SY, Piers E, Bohlmann J. Loblolly pine abietadienol/abietadienal oxidase PtAO (CYP720B1) is a multifunctional, multisubstrate cytochrome P450 monooxygenase. Proc Natl Acad Sci U S A 102(22):8060-5. (2005).

Lippert D, Zhuang J, Ralph S, Ellis DE, Gilbert M, Olafson R, Ritland K, Ellis B, Douglas CJ, Bohlmann J. Proteome analysis of early somatic embryogenesis in Picea glauca. Proteomics 5(2):461-73. (2005).

Miller B, Madilao LL, Ralph S, Bohlmann J. Insect-induced conifer defense. White pine weevil and methyl jasmonate induce traumatic resinosis, de novo formed volatile emissions, and accumulation of terpenoid synthase and putative octadecanoid pathway transcripts in Sitka spruce. Plant Physiol 137(1):369-82. (2005).

Martin D and Bohlmann J Molecular biochemistry and genomics of terpenoid defenses in conifers. Recent Advances in Phytochemistry (39):29-56. (2005).

Rungis D, Hamberger B, Berube Y, Wilkin J, Bohlmann J, and K Ritland Efficient genetic mapping of single nucleotide polymorphisms based upon DNA mismatch digestion. Molecular Breeding (16):261-270. (2005).

Huber DP, Ralph S, Bohlmann J. Genomic hardwiring and phenotypic plasticity of terpenoid-based defenses in conifers. J Chem Ecol 30(12):2399-418. (2004).

Rungis D, Berube Y, Zhang J, Ralph S, Ritland CE, Ellis BE, Douglas C, Bohlmann J, Ritland K. Robust simple sequence repeat markers for spruce (Picea spp.) from expressed sequence tags. Theor Appl Genet 109(6):1283-94. (2004).

Lucker J, Bowen P, Bohlmann J. Vitis vinifera terpenoid cyclases: functional identification of two sesquiterpene synthase cDNAs encoding (+)-valencene synthase and (-)-germacrene D synthase and expression of mono- and sesquiterpene synthases in grapevine flowers and berries. Phytochemistry 65(19):2649-59. (2004).

Chen F, Ro DK, Petri J, Gershenzon J, Bohlmann J, Pichersky E, Tholl D. Characterization of a root-specific Arabidopsis terpene synthase responsible for the formation of the volatile monoterpene 1,8-cineole. Plant Physiol 135(4):1956-66. (2004).

Arimura G, Ozawa R, Kugimiya S, Takabayashi J, Bohlmann J. Herbivore-induced defense response in a model legume. Two-spotted spider mites induce emission of (E)-beta-ocimene and transcript accumulation of (E)-beta-ocimene synthase in Lotus japonicus. Plant Physiol 135(4):1976-83. (2004).

Martin DM, Fäldt J, Bohlmann J. Functional characterization of nine Norway Spruce TPS genes and evolution of gymnosperm terpene synthases of the TPS-d subfamily. Plant Physiol 135(4):1908-27. (2004).

Reiling KK, Yoshikuni Y, Martin VJ, Newman J, Bohlmann J, Keasling JD. Mono and diterpene production in Escherichia coli. Biotechnol Bioeng 87(2):200-12. (2004).

Kolosova N, Miller B, Ralph S, Ellis BE, Douglas C, Ritland K, Bohlmann J. Isolation of high-quality RNA from gymnosperm and angiosperm trees. Biotechniques 36(5):821-4. (2004).

Martin DM, Bohlmann J. Identification of Vitis vinifera (-)-alpha-terpineol synthase by in silico screening of full-length cDNA ESTs and functional characterization of recombinant terpene synthase. Phytochemistry 65(9):1223-9. (2004).

Arimura G, Huber DP, Bohlmann J. Forest tent caterpillars (Malacosoma disstria) induce local and systemic diurnal emissions of terpenoid volatiles in hybrid poplar (Populus trichocarpa x deltoides): cDNA cloning, functional characterization, and patterns of gene expression of (-)-germacrene D synthase, PtdTPS1. Plant J 37(4):603-16. (2004).

Huber DPW and J Bohlmann Terpene synthases and the mediation of plant-insect ecological interactions by terpenoids: a mini-review. In: Plant Adaptation: Molecular Genetics and Ecology. Q.C.B. Cronck, J. Whitton, R.H. Ree, I.E.P. Taylor. NRC Research Press, Ottawa, Ontario 70 – 81. (2004).

McKay SA, Hunter WL, Godard KA, Wang SX, Martin DM, Bohlmann J, Plant AL. Insect attack and wounding induce traumatic resin duct development and gene expression of (-)-pinene synthase in Sitka spruce. Plant Physiol 133(1):368-78. (2003).

Martin DM, Gershenzon J, Bohlmann J. Induction of volatile terpene biosynthesis and diurnal emission by methyl jasmonate in foliage of Norway spruce. Plant Physiol 132(3):1586-99. (2003).

Dudareva N, Martin D, Kish CM, Kolosova N, Gorenstein N, Fäldt J, Miller B, Bohlmann J. (E)-beta-ocimene and myrcene synthase genes of floral scent biosynthesis in snapdragon: function and expression of three terpene synthase genes of a new terpene synthase subfamily. Plant Cell 15(5):1227-41. (2003).

Martin D, Bohlmann J, Gershenzon J, Francke W, Seybold SJ. A novel sex-specific and inducible monoterpene synthase activity associated with a pine bark beetle, the pine engraver, Ips pini. Naturwissenschaften 90(4):173-9. (2003).

Fäldt J, Arimura G, Gershenzon J, Takabayashi J, Bohlmann J. Functional identification of AtTPS03 as (E)-beta-ocimene synthase: a monoterpene synthase catalyzing jasmonate- and wound-induced volatile formation in Arabidopsis thaliana. Planta 216(5):745-51. (2003).

Fäldt J, Martin D, Miller B, Rawat S, Bohlmann J. Traumatic resin defense in Norway spruce (Picea abies): methyl jasmonate-induced terpene synthase gene expression, and cDNA cloning and functional characterization of (+)-3-carene synthase. Plant Mol Biol 51(1):119-33. (2003).

Aubourg S, Lecharny A, Bohlmann J. Genomic analysis of the terpenoid synthase ( AtTPS) gene family of Arabidopsis thaliana. Mol Genet Genomics 267(6):730-45. (2002).

Martin D, Tholl D, Gershenzon J, Bohlmann J. Methyl jasmonate induces traumatic resin ducts, terpenoid resin biosynthesis, and terpenoid accumulation in developing xylem of Norway spruce stems. Plant Physiol 129(3):1003-18. (2002).

Bohlmann J, Stauber EJ, Krock B, Oldham NJ, Gershenzon J, Baldwin IT. Gene expression of 5-epi-aristolochene synthase and formation of capsidiol in roots of Nicotiana attenuata and N. sylvestris. Phytochemistry 60(2):109-16. (2002).

Alfaro RI, Borden JH, King JN, Tomlin ES, McIntosh RL, and J Bohlmann Mechanisms of Resistance in Conifers against shoot infesting insects. In: "Mechanisms and Deployment of Resistance in Trees to Insects." M.R. Wagner, K.M. Clancy, F. Lieutier, and T.D. Paine. Kluwer Academic Press, Dordrecht, the Netherlands 101-126. (2002).