-
[show abstract]
[hide abstract]
ABSTRACT: Sexual reproduction in fungi is regulated by the mating-type () locus where recombination is suppressed. We investigated the evolution of loci in eight fungal species belonging to and (Sordariomycetes, Ascomycota) that include conifer pathogens and beetle symbionts. The idiomorph/allele was identified from the assembled and annotated genome, and the locus is flanked by genes coding for cytoskeleton protein () and DNA lyase. The synteny of these genes is conserved and consistent with other members in Ascomycota. Using sequences from and flanking regions, we characterized the idiomorph from other isolates of and performed dotplot analysis between the two idiomorphs. Unexpectedly, the idiomorph contains a truncated gene upstream of the gene that bears the high-mobility-group domain. The nucleotide and amino acid sequence of the truncated gene is similar to its homologous copy in the idiomorph in the opposite mating-type isolate, except that positive selection is acting on the truncated gene and the alpha(α)-box that encodes the transcription factor has been deleted. The idiomorphs sharing identical gene organization were present in seven additional species in the Ophiostomatales, suggesting that the presence of truncated gene is a general pattern in this order. We propose that an ancient unequal recombination event resulted in the ancestral gene integrated into the idiomorph and surviving as the truncated genes. The α-box domain of gene, located at the same locus adjacent to the gene, could have been removed by deletion after recombination due to mating signal interference. Our data confirmed a 1:1 MAT/sex ratio in two pathogen populations, and showed that all members of the Ophiostomatales studied here including those that were previously deemed asexual have the potential to reproduce sexually. This ability can potentially increase genetic variability and can enhance fitness in new, ecological niches.
G3 (Bethesda, Md.). 03/2013; 3(3):465-80.
-
[show abstract]
[hide abstract]
ABSTRACT: Diterpene resin acids (DRAs) are major components of pine (Pinus spp.) oleoresin. They play critical roles in conifer defense against insects and pathogens and as a renewable resource for industrial bioproducts. The core structures of DRAs are formed in secondary (i.e. specialized) metabolism via cycloisomerization of geranylgeranyl diphosphate (GGPP) by diterpene synthases (diTPSs). Previously described gymnosperm diTPSs of DRA biosynthesis are bifunctional enzymes that catalyze the initial bicyclization of GGPP followed by rearrangement of a (+)-copalyl diphosphate intermediate at two discrete class II and class I active sites. In contrast, similar diterpenes of gibberellin primary (i.e. general) metabolism are produced by the consecutive activity of two monofunctional class II and class I diTPSs. Using high-throughput transcriptome sequencing, we discovered 11 diTPS from jack pine (Pinus banksiana) and lodgepole pine (Pinus contorta). Three of these were orthologous to known conifer bifunctional levopimaradiene/abietadiene synthases. Surprisingly, two sets of orthologous PbdiTPSs and PcdiTPSs were monofunctional class I enzymes that lacked functional class II active sites and converted (+)-copalyl diphosphate, but not GGPP, into isopimaradiene and pimaradiene as major products. Diterpene profiles and transcriptome sequences of lodgepole pine and jack pine are consistent with roles for these diTPSs in DRA biosynthesis. The monofunctional class I diTPSs of DRA biosynthesis form a new clade within the gymnosperm-specific TPS-d3 subfamily that evolved from bifunctional diTPS rather than monofunctional enzymes (TPS-c and TPS-e) of gibberellin metabolism. Homology modeling suggested alterations in the class I active site that may have contributed to their functional specialization relative to other conifer diTPSs.
Plant physiology 02/2013; 161(2):600-616. · 6.53 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Grosmannia clavigera is a bark beetle-vectored pine pathogen in the mountain pine beetle epidemic in western North America. Grosmannia clavigera colonizes pines despite the trees' massive oleoresin terpenoid defences. We are using a functional genomics approach to identify G. clavigera's mechanisms of adaptation to pine defences. We annotated the ABC transporters in the G. clavigera genome and generated RNA-seq transcriptomes from G. clavigera grown with a range of terpenes. We functionally characterized GcABC-G1, a pleiotropic drug resistance (PDR) transporter that was highly induced by terpenes, using qRT-PCR, gene knock-out and heterologous expression in yeast. Deleting GcABC-G1 increased G. clavigera's sensitivity to monoterpenes and delayed development of symptoms in inoculated young lodgepole pine trees. Heterologous expression of GcABC-G1 in yeast increased tolerance to monoterpenes. G. clavigera but not the deletion mutant, can use (+)-limonene as a carbon source. Phylogenetic analysis placed GcABC-G1 outside the ascomycete PDR transporter clades. G. clavigera appears to have evolved two mechanisms to survive and grow when exposed to monoterpenes: GcABC-G1 controls monoterpene levels within the fungal cells and G. clavigera uses monoterpenes as a carbon source. This work has implications for understanding adaptation to host defences in an important forest insect-fungal system, and potentially for metabolic engineering of terpenoid production in yeast.
New Phytologist 12/2012; · 6.64 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: BACKGROUND: Mountain pine beetles, Dendroctonus ponderosae Hopkins (Coleoptera: Curculionidae), are native to western North America, but have recently begun to expand their range across the Canadian Rocky Mountains. The requirement for larvae to withstand extremely cold winter temperatures and potentially toxic host secondary metabolites in the midst of their ongoing development makes this a critical period of their lives. RESULTS: We have uncovered global protein profiles for overwintering mountain pine beetle larvae. We have also quantitatively compared the proteomes for overwintering larvae sampled during autumn cooling and spring warming using iTRAQ methods. We identified 1507 unique proteins across all samples. In total, 33 proteins exhibited differential expression (FDR < 0.05) when compared between larvae before and after a cold snap in the autumn; and 473 proteins exhibited differential expression in the spring when measured before and after a steady incline in mean daily temperature. Eighteen proteins showed significant changes in both autumn and spring samples. CONCLUSIONS: These first proteomic data for mountain pine beetle larvae show evidence of the involvement of trehalose, 2-deoxyglucose, and antioxidant enzymes in overwintering physiology; confirm and expand upon previous work implicating glycerol in cold tolerance in this insect; and provide new, detailed information on developmental processes in beetles. These results and associated data will be an invaluable resource for future targeted research on cold tolerance mechanisms in the mountain pine beetle and developmental biology in coleopterans.
Insect biochemistry and molecular biology 09/2012; · 3.25 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: In developing grapevine (Vitis vinifera L.) berries, precursor volatile organic compounds (PVOCs) are largely stored as glycosides which may be hydrolyzed to release VOCs during fruit ripening, wine making, or aging. VOCs can be further transformed by yeast metabolism. Together, these processes contribute to complexity of wine aromas. Floral and citrus odors of many white wine varietals are attributed to monoterpenes and monoterpene alcohols, while phenolic compounds, norisoprenoids, and other volatiles also play important roles in determining aroma. We present an analysis of PVOCs stored as glycosides in developing Gewürztraminer berries during the growing season. We optimized a method for PVOC analysis suitable for small amounts of Muscat grapevine berries and showed that the amount of PVOCs dramatically increased during and after véraison. Transcript profiling of the same berry samples underscored the involvement of terpenoid pathway genes in the accumulation of PVOCs. The onset of monoterpenol PVOC accumulation in developing grapes was correlated with an increase of transcript abundances of early terpenoid pathway enzymes. Transcripts encoding the methylerythritol phosphate pathway gene 4-hydroxy-3-methylbut-2-enyl diphosphate reductase, as well as geraniol diphosphate synthase, were up-regulated preceding and during the increase in monoterpenol PVOCs. Transcripts for linalool/nerolidol synthase increased in later véraison stages.
Planta 07/2012; 236(3):919-29. · 3.00 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The mountain pine beetle (MPB) is a native bark beetle of western North America that attacks pine tree species, particularly lodgepole pine. It is closely associated with the ophiostomatoid ascomycetes Grosmannia clavigera, Leptographium longiclavatum, Ophiostoma montium, and Ceratocystiopsis sp.1, with which it is symbiotically associated. To develop a better understanding of interactions between beetles, fungi, and host trees, we used target-specific DNA primers with qPCR to assess the changes in fungal associate abundance over the stages of the MPB life cycle that occur in galleries under the bark of pine trees. Multivariate analysis of covariance identified statistically significant changes in the relative abundance of the fungi over the life cycle of the MPB. Univariate analysis of covariance identified a statistically significant increase in the abundance of Ceratocystiopsis sp.1 through the beetle life cycle, and pair-wise analysis showed that this increase occurs after the larval stage. In contrast, the abundance of O. montium and Leptographium species (G. clavigera, L. longiclavatum) did not change significantly through the MPB life cycle. From these results, the only fungus showing a significant increase in relative abundance has not been formally described and has been largely ignored by other MPB studies. Although our results were from only one site, in previous studies we have shown that the fungi described were all present in at least ten sites in British Columbia. We suggest that the role of Ceratocystiopsis sp.1 in the MPB system should be explored, particularly its potential as a source of nutrients for teneral adults.
Microbial Ecology 06/2012; 64(4):909-17. · 2.91 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Proteomics research is hampered in many organisms due to a lack of an appropriate reference genome sequence that can be used
in the interpretation of tandem mass spectrometry data for the identification of proteins. Public DNA sequence repositories
have grown to considerable size and can, in most cases, serve to provide at least partial interpretation of a large-scale
proteomics dataset. However, when species-specific sequences or sequences from a closely related species are available, a
boutique sequence database can provide considerable increases in specificity, confidence, and completeness of protein identification.
Here, we describe the development of a protein database from a large-scale expressed sequence tag and full-length complementary
DNA sequencing project in the economically and ecologically important spruce (Picea) genus.
Tree Genetics & Genomes 04/2012; 5(4):723-727. · 2.34 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: A collection of cDNA libraries from white spruce (Picea glauca) and interior spruce (P. glauca × engelmanii) vascular tissue were analyzed to identify a set of genes that could serve as tissue-related markers within the coniferous
vascular system. Multivariate exploratory methods identified up to 128 genes co-expressed similarly among three xylem libraries.
The majority (87) of these genes formed three distinctive meta-clusters, denoting putative gene cliques in xylem tissue. Of
the selected genes, 33 (25%) exhibited no significant sequence homology in queries against any public databases, indicating
the possibility of their unique expression in the xylem tissue of conifers. Another 38 genes (30%) had ambiguous annotation.
Validation of the annotated genes with analog data, obtained from a wet-lab study utilizing microarray slides with 18,881
spots, resulted in a screened list of 29 genes as xylem-related markers. Response to stress was the predominant category to
which the screened genes corresponded. Among the screened genes, elements of the phenolics biosynthesis, cinnamyl alcohol
dehydrogenase and laccase, together with the fundamental enzyme of the cell wall biosynthesis, cellulose synthase, prominently
delineated characteristics of the wood-forming tissue, xylem.
KeywordsConifer–EST–Digital gene expression–Spruce–Vascular tissue–Xylem
Tree Genetics & Genomes 04/2012; 6(4):601-611. · 2.34 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Conifers have evolved elaborate inducible, terpenoid-based defense mechanisms to deter attacks from bark beetles and other herbivore species. Herbivore damage triggers the production of oleoresin containing monoterpene, sesquiterpene and diterpene components that serve as toxins and physical barriers to herbivore invasion. Induced terpene formation appears to be regulated by specific enzymes of terpene metabolism whose activity increases on herbivore damage. Among the best studied of these are terpene synthases, enzymes which convert acyclic prenyl diphosphates to the parent terpene skeletons. Terpene synthase activity in turn is regulated by the transcription of terpene synthase genes. Induced terpene biosynthesis is also often accompanied by extensive cellular differentiation, including the formation of new resin ducts. The signal transduction cascades that initiate these shifts in conifer metabolism and cell differentiation are poorly understood due to the lack of well-developed model systems and appropriate genetic mutants. However, there are strong indications that octadecanoid pathway metabolites and ethylene have roles in this signaling, as they do in defense signaling in angiosperms. There are still large gaps in our knowledge of the signal transduction networks leading to herbivore-induced terpenoid accumulation in conifers. However, the development of new genomic, proteomic and metabolomic tools, as well as the establishment of convenient in vitro systems should facilitate more rapid advances in this field in the near future. The results will have important implications for understanding the evolution of conifer defense mechanisms as well as for the management of commercially important forest tree species, such as spruce, pine, and fir.
Phytochemistry Reviews 04/2012; 5(1):179-189. · 4.33 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The use of single-nucleotide polymorphisms
(SNPs) as molecular markers in plant studies has become
increasingly common. With the development of these
markers, there is an interest in determining levels of variation
in natural populations. Here, we identify and characterize
levels of SNPs in wild populations of aspen (Populus tremuloides
Michx.). Four populations were sampled from Alberta
and British Columbia in Western Canada. A total of 35 gene
regions were selected for analysis. The loci selected are mainly
involved in wood formation and include regions from genes
for lignin biosynthesis, cellulose biosynthesis, and other cell
wall compounds and a number of transcription factors. Other
genes included those coding for growth hormones, disease
resistance, and light responses. Primers were developed from
conserved regions in multi-species EST alignments. Regions
were PCR amplified, and products (approximately 500 to
1,000 bp) were assessed for levels of SNPs using Ecotilling.
From a total of approximately 25 kb 462 SNPs were identified,
over 18 SNPs/kb. Thus, SNPs are an abundant and
potentially useful molecular marker in aspen. Gene diversity
(heterozygosity) varied in the gene regions, with an overall
average of HT00.18. Although gene diversity was considerable,
genetic differentiation was low with the overall FST
value being 0.004. A surrogate measure of linkage disequilibrium
(LD) was calculated, and overall, the LD was shown to
decay relatively rapidly with distance along the gene region.
The results obtained from the wood formation genes in this
study will enable further targeting of regions for association
studies on lignin and cellulose variation in aspen and other
Populus species.
Tree Genetics & Genomes 04/2012; · 2.34 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The labdanoid diterpene alcohol cis-abienol is a major component of the aromatic oleoresin of balsam fir (Abies balsamea) and serves as a valuable bioproduct material for the fragrance industry. Using high-throughput 454 transcriptome sequencing and metabolite profiling of balsam fir bark tissue, we identified candidate diterpene synthase sequences for full-length cDNA cloning and functional characterization. We discovered a bifunctional class I/II cis-abienol synthase (AbCAS), along with the paralogous levopimaradiene/abietadiene synthase and isopimaradiene synthase, all of which are members of the gymnosperm-specific TPS-d subfamily. The AbCAS-catalyzed formation of cis-abienol proceeds via cyclization and hydroxylation at carbon C-8 of a postulated carbocation intermediate in the class II active site, followed by cleavage of the diphosphate group and termination of the reaction sequence without further cyclization in the class I active site. This reaction mechanism is distinct from that of synthases of the isopimaradiene- or levopimaradiene/abietadiene synthase type, which employ deprotonation reactions in the class II active site and secondary cyclizations in the class I active site, leading to tricyclic diterpenes. Comparative homology modeling suggested the active site residues Asp-348, Leu-617, Phe-696, and Gly-723 as potentially important for the specificity of AbCAS. As a class I/II bifunctional enzyme, AbCAS is a promising target for metabolic engineering of cis-abienol production.
Journal of Biological Chemistry 02/2012; 287(15):12121-31. · 4.77 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Conifer diterpene synthases (diTPSs) catalyze the multi-step cycloisomerization of geranylgeranyl diphosphate, or copalyl diphosphate, to a variety of diterpenes in general (i.e., primary) and specialized (i.e., secondary) metabolism. Despite their functional diversity, the known conifer diTPSs are structurally closely related, with variations in three conserved domains, α, β and γ. The catalytic specificity of conifer class I and class I/II diTPSs is predominantly determined by the protein environment of the C-terminal class I active site through stabilization of common and unique carbocation intermediates. Using the crystal structure of Taxus brevifolia taxadiene synthase as template, comparative modeling and mutagenesis of the class I diTPS ent-kaurene synthase from Picea glauca (PgKS) was performed to elucidate the catalytic specificity of PgKS relative to spruce diTPSs of specialized metabolism. N-terminal truncations demonstrated a role for the βγ domain in class I enzyme activity for PgKS, facilitating the closure of the class I active site upon substrate binding. Based on position, Arg476 and Asp736 in the C-terminal α domain of PgKS may contribute to this conformational transition and appear critical for catalysis. Consistent with the mechanism of other diTPSs, the subsequent ionization of a copalyl diphosphate substrate and coordination of the diphosphate group is controlled by strictly conserved residues in the DDxxD and NDIQGCKRE motif of PgKS, such as Asn656 and Arg653. Furthermore, Lys478, Trp502, Met588, Ala615 and Ile619 control the enzymatic activity and specificity of PgKS via carbocation stabilization en route to ent-kaurene. These positions show a high level of amino acid variation, consistent with functional plasticity among conifer diTPSs of different functions in general or specialized metabolism.
Phytochemistry 12/2011; 74:30-9. · 3.35 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Diterpene resin acids (DRAs) are specialized (secondary) metabolites of the oleoresin defense of conifers produced by diterpene synthases and cytochrome P450s of the CYP720B family. The evolution of DRA metabolism shares common origins with the biosynthesis of ent-kaurenoic acid, which is highly conserved in general (primary) metabolism of gibberellin biosynthesis. Transcriptome mining in species of spruce (Picea) and pine (Pinus) revealed CYP720Bs of four distinct clades. We cloned a comprehensive set of 12 different Sitka spruce (Picea sitchensis) CYP720Bs as full-length cDNAs. Spatial expression profiles, methyl jasmonate induction, and transcript enrichment in terpenoid-producing resin ducts suggested a role of CYP720B4 in DRA biosynthesis. CYP720B4 was characterized as a multisubstrate, multifunctional enzyme by the formation of oxygenated diterpenoids in metabolically engineered yeast, yeast in vivo transformation of diterpene substrates, in vitro assays with CYP720B4 protein produced in Escherichia coli, and alteration of DRA profiles in RNA interference-suppressed spruce seedlings. CYP720B4 was active with 24 different diterpenoid substrates, catalyzing consecutive C-18 oxidations in the biosynthesis of an array of diterpene alcohols, aldehydes, and acids. CYP720B4 was most active in the formation of dehydroabietic acid, a compound associated with insect resistance of Sitka spruce. We identified patterns of convergent evolution of CYP720B4 in DRA metabolism and ent-kaurene oxidase CYP701 in gibberellin metabolism and revealed differences in the evolution of specialized and general diterpene metabolism in a gymnosperm. The genomic and functional characterization of the gymnosperm CYP720B family highlights that the evolution of specialized metabolism involves substantial diversification relative to conserved, general metabolism.
Plant physiology 12/2011; 157(4):1677-95. · 6.53 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: We investigated the population structure of Grosmannia clavigera (Gc), a fungal symbiont of the mountain pine beetle (MPB) that plays a crucial role in the establishment and reproductive success of this pathogen. This insect-fungal complex has destroyed over 16 million ha of lodgepole pine forests in Canada, the largest MPB epidemic in recorded history. During this current epidemic, MPB has expanded its range beyond historically recorded boundaries, both northward and eastward, and has now reached the jack pine of Alberta, potentially threatening the Canadian boreal forest. To better understand the dynamics between the beetle and its fungal symbiont, we sampled 19 populations in western North America and genotyped individuals from these populations with eight microsatellite markers. The fungus displayed high haplotype diversity, with over 250 unique haplotypes observed in 335 single spore isolates. Linkage equilibria in 13 of the 19 populations suggested that the fungus reproduces sexually. Bayesian clustering and distance analyses identified four genetic clusters that corresponded to four major geographical regions, which suggested that the epidemic arose from multiple geographical sources. A genetic cluster north of the Rocky Mountains, where the MPB has recently become established, experienced a population bottleneck, probably as a result of the recent range expansion. The two genetic clusters located north and west of the Rocky Mountains contained many fungal isolates admixed from all populations, possibly due to the massive movement of MPB during the epidemic. The general agreement in north-south differentiation of MPB and G. clavigera populations points to the fungal pathogen's dependence on the movement of its insect vector. In addition, the patterns of diversity and the individual assignment tests of the fungal associate suggest that migration across the Rocky Mountains occurred via a northeastern corridor, in accordance with meteorological patterns and observation of MPB movement data. Our results highlight the potential of this pathogen for both expansion and sexual reproduction, and also identify some possible barriers to gene flow. Understanding the ecological and evolutionary dynamics of this fungus-beetle association is important for the modelling and prediction of MPB epidemics.
Molecular Ecology 11/2011; 21(1):71-86. · 5.52 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: The levopimaradiene/abietadiene synthase from Norway spruce (Picea abies; PaLAS) has previously been reported to produce a mixture of four diterpene hydrocarbons when incubated with geranylgeranyl diphosphate as the substrate: levopimaradiene, abietadiene, neoabietadiene, and palustradiene. However, variability in the assay products observed by GC-MS of this and orthologous conifer diterpene synthases over the past 15 years suggested that these diterpenes may not be the initial enzyme assay products but are rather the products of dehydration of an unstable alcohol. We have identified epimers of the thermally unstable allylic tertiary alcohol 13-hydroxy-8(14)-abietene as the products of PaLAS. The identification of these compounds, not previously described in conifers, as the initial products of PaLAS has considerable implications for our understanding of the complexity of the biosynthetic pathway of the structurally diverse diterpene resin acids of conifer defense.
Journal of Biological Chemistry 06/2011; 286(24):21145-53. · 4.77 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Grosmannia clavigera is a fungal pathogen of pine forests in western North America and a symbiotic associate of two sister bark beetles: Dendroctonus ponderosae and D. jeffreyi. This fungus and its beetle associate D. ponderosae are expanding in large epidemics in western North America. Using the fungal genome sequence and gene annotations, we assessed whether fungal isolates from the two beetles inhabiting different species of pine in epidemic regions of western Canada and the USA, as well as in localized populations outside of the current epidemic, represent different genetic lineages. We characterized nucleotide variations in 67 genomic regions and selected 15 for the phylogenetic analysis. Using concordance of gene genealogies and distinct ecological characteristics, we identified two sibling phylogenetic species: Gc and Gs. Where the closely related Pinus ponderosa and P. jeffreyi are infested by localized populations of their respective beetles, Gc is present. In contrast, Gs is an exclusive associate of D. ponderosae mainly present on its primary host-tree P. contorta; however, in the current epidemic areas, it is also found in other pine species. These results suggest that the host-tree species and the beetle population dynamics may be important factors associated with the genetic divergence and diversity of fungal partners in the beetle-tree ecosystems. Gc represents the original G. clavigera holotype, and Gs should be described as a new species.
Molecular Ecology 06/2011; 20(12):2581-602. · 5.52 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Some plant terpenes such as sterols and carotenes are part of primary metabolism and found essentially in all plants. However, the majority of the terpenes found in plants are classified as 'secondary' compounds, those chemicals whose synthesis has evolved in plants as a result of selection for increased fitness via better adaptation to the local ecological niche of each species. Thousands of such terpenes have been found in the plant kingdom, but each species is capable of synthesizing only a small fraction of this total. In plants, a family of terpene synthases (TPSs) is responsible for the synthesis of the various terpene molecules from two isomeric 5-carbon precursor 'building blocks', leading to 5-carbon isoprene, 10-carbon monoterpenes, 15-carbon sesquiterpenes and 20-carbon diterpenes. The bryophyte Physcomitrella patens has a single TPS gene, copalyl synthase/kaurene synthase (CPS/KS), encoding a bifunctional enzyme producing ent-kaurene, which is a precursor of gibberellins. The genome of the lycophyte Selaginella moellendorffii contains 18 TPS genes, and the genomes of some model angiosperms and gymnosperms contain 40-152 TPS genes, not all of them functional and most of the functional ones having lost activity in either the CPS- or KS-type domains. TPS genes are generally divided into seven clades, with some plant lineages having a majority of their TPS genes in one or two clades, indicating lineage-specific expansion of specific types of genes. Evolutionary plasticity is evident in the TPS family, with closely related enzymes differing in their product profiles, subcellular localization, or the in planta substrates they use.
The Plant Journal 04/2011; 66(1):212-29. · 6.16 Impact Factor
-
Kiyokazu Agata,
Samer Alasaad,
Vera Maria Fonseca Almeida-Val,
J A Alvarez-Dios,
F Barbisan,
Jon S Beadell,
J F Beltrán,
M Benítez,
G Bino,
Colin Bleay, [......],
Myriam Valenzuela-Bustamante,
M S Van de Vliet,
K Vandepitte,
M Vera,
Peter Wandeler,
Weimin Wang,
Yong-Jin Won,
A Yamashiro,
T Yamashiro,
Changcheng Zhu
[show abstract]
[hide abstract]
ABSTRACT: This article documents the addition of 238 microsatellite marker loci to the Molecular Ecology Resources Database. Loci were developed for the following species: Alytes dickhilleni, Arapaima gigas, Austropotamobius italicus, Blumeria graminis f. sp. tritici, Cobitis lutheri, Dendroctonus ponderosae, Glossina morsitans morsitans, Haplophilus subterraneus, Kirengeshoma palmata, Lysimachia japonica, Macrolophus pygmaeus, Microtus cabrerae, Mytilus galloprovincialis, Pallisentis (Neosentis) celatus, Pulmonaria officinalis, Salminus franciscanus, Thais chocolata and Zootoca vivipara. These loci were cross-tested on the following species: Acanthina monodon, Alytes cisternasii, Alytes maurus, Alytes muletensis, Alytes obstetricans almogavarii, Alytes obstetricans boscai, Alytes obstetricans obstetricans, Alytes obstetricans pertinax, Cambarellus montezumae, Cambarellus zempoalensis, Chorus giganteus, Cobitis tetralineata, Glossina fuscipes fuscipes, Glossina pallidipes, Lysimachia japonica var. japonica, Lysimachia japonica var. minutissima, Orconectes virilis, Pacifastacus leniusculus, Procambarus clarkii, Salminus brasiliensis and Salminus hilarii.
Molecular Ecology Resources 04/2011; 11(3):586-9. · 3.06 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: In conifers, terpene synthases (TPSs) of the gymnosperm-specific TPS-d subfamily form a diverse array of mono-, sesqui-, and diterpenoid compounds, which are components of the oleoresin secretions and volatile emissions. These compounds contribute to defence against herbivores and pathogens and perhaps also protect against abiotic stress.
The availability of extensive transcriptome resources in the form of expressed sequence tags (ESTs) and full-length cDNAs in several spruce (Picea) species allowed us to estimate that a conifer genome contains at least 69 unique and transcriptionally active TPS genes. This number is comparable to the number of TPSs found in any of the sequenced and well-annotated angiosperm genomes. We functionally characterized a total of 21 spruce TPSs: 12 from Sitka spruce (P. sitchensis), 5 from white spruce (P. glauca), and 4 from hybrid white spruce (P. glauca × P. engelmannii), which included 15 monoterpene synthases, 4 sesquiterpene synthases, and 2 diterpene synthases.
The functional diversity of these characterized TPSs parallels the diversity of terpenoids found in the oleoresin and volatile emissions of Sitka spruce and provides a context for understanding this chemical diversity at the molecular and mechanistic levels. The comparative characterization of Sitka spruce and Norway spruce diterpene synthases revealed the natural occurrence of TPS sequence variants between closely related spruce species, confirming a previous prediction from site-directed mutagenesis and modelling.
BMC Plant Biology 03/2011; 11:43. · 3.45 Impact Factor
-
[show abstract]
[hide abstract]
ABSTRACT: Sandalwood oil is one of the worlds most highly prized fragrances. To identify the genes and encoded enzymes responsible for santalene biosynthesis, we cloned and characterized three orthologous terpene synthase (TPS) genes SaSSy, SauSSy, and SspiSSy from three divergent sandalwood species; Santalum album, S. austrocaledonicum, and S. spicatum, respectively. The encoded enzymes catalyze the formation of α-, β-, epi-β-santalene, and α-exo-bergamotene from (E,E)-farnesyl diphosphate (E,E-FPP). Recombinant SaSSy was additionally tested with (Z,Z)-farnesyl diphosphate (Z,Z-FPP) and remarkably, found to produce a mixture of α-endo-bergamotene, α-santalene, (Z)-β-farnesene, epi-β-santalene, and β-santalene. Additional cDNAs that encode bisabolene/bisabolol synthases were also cloned and functionally characterized from these three species. Both the santalene synthases and the bisabolene/bisabolol synthases reside in the TPS-b phylogenetic clade, which is more commonly associated with angiosperm monoterpene synthases. An orthologous set of TPS-a synthases responsible for formation of macrocyclic and bicyclic sesquiterpenes were characterized. Strict functionality and limited sequence divergence in the santalene and bisabolene synthases are in contrast to the TPS-a synthases, suggesting these compounds have played a significant role in the evolution of the Santalum genus.
Journal of Biological Chemistry 03/2011; 286(20):17445-54. · 4.77 Impact Factor
