tremulacin and salicortin

The survival of insect herbivores on chemically defended plants may often depend on their ability to metabolize these defense compounds. However, only little knowledge is available on how insects actually process most plant defense compounds. We investigated the metabolism of salicinoids, a major group of phenolic glycosides in poplar and willow species, by a generalist herbivore, the gypsy moth (Lymantria dispar). Seven salicinoid metabolites identified in gypsy moth caterpillar feces were mostly conjugates with glucose, cysteine or glycine. Two of the glucosides were phosphorylated, a feature not previously reported for insect metabolites of plant defense compounds. The origins of these metabolites were traced to specific moieties of three major poplar salicinoids ingested, salicin, salicortin and tremulacin. Based on the observed metabolite patterns we were able to deduce the initial steps of salicinoid breakdown in L. dispar guts, which involves cleavage of ester bonds. The conjugated molecules were effectively eliminated within 24 h after ingestion. Some of the initial breakdown products (salicin and catechol) demonstrated negative effects on insect growth and survival in bioassays on artificial diets. Gypsy moth caterpillars with prior feeding experience on salicinoid-containing poplar foliage converted salicinoids to the identified metabolites more efficiently than caterpillars pre-fed an artificial diet. The majority of the metabolites we identified were also produced by other common poplar-feeding insects. The conversion of plant defenses like salicinoids to a variety of water-soluble sugar, phosphate and amino acid conjugates and their subsequent excretion fits the general detoxification strategy found in insect herbivores and other animals.

Topics: Animals; Benzyl Alcohols; Feeding Behavior; Glucosides; Herbivory; Larva; Moths; Plant Leaves; Populus; Tissue Distribution

The absolute configuration of salicortin, HCH-salicortin and tremulacin, isolated from leaves of Populus trichocarpa × deltoides Beaupré, was determined by comparing spectroscopic data of these compounds with those of idescarpin, isolated from leaves of Idesia polycarpa. All compounds were characterized by nuclear magnetic resonance spectroscopy, high-resolution mass spectrometry, and circular dichroism spectroscopy. It was found that the hydroxy cyclohexenonoyl (HCH) moiety in all compounds is (S)-configured. In addition, it was shown that leaves of Idesia polycarpa contain high amounts of (-)-idescarpin (1.1%, based on dry weight).

Topics: Circular Dichroism; Cyclohexanecarboxylic Acids; Glucosides; Mass Spectrometry; Molecular Structure; Plant Leaves; Populus; Salicaceae

Salicinoids are phenolic glycosides (PGs) characteristic of the Salicaceae and are known defenses against insect herbivory. Common examples are salicin, salicortin, tremuloidin, and tremulacin, which accumulate to high concentrations in the leaves and bark of willows and poplars. Although their biosynthetic pathway is not known, recent work has suggested that benzyl benzoate may be a potential biosynthetic intermediate. Two candidate genes, named PtACT47 and PtACT49, encoding BAHD-type acyl transferases were identified and are predicted to produce such benzylated secondary metabolites. Herein described are the cDNA cloning, heterologous expression and in vitro functional characterization of these two BAHD acyltransferases. Recombinant PtACT47 exhibited low substrate selectivity and could utilize acetyl-CoA, benzoyl-CoA, and cinnamoyl-CoA as acyl donors with a variety of alcohols as acyl acceptors. This enzyme showed the greatest Km/Kcat ratio (45.8 nM(-1) s(-1)) and lowest Km values (45.1 μM) with benzoyl-CoA and salicyl alcohol, and was named benzoyl-CoA: salicyl alcohol O-benzoyltransferase (PtSABT). Recombinant PtACT49 utilized a narrower range of substrates, including benzoyl-CoA and acetyl-CoA and a limited number of alcohols. Its highest Km/Kcat (31.8 nM(-1) s(-1)) and lowest Km (55.3 μM) were observed for benzoyl-CoA and benzyl alcohol, and it was named benzoyl-CoA: benzyl alcohol O-benzoyltransferase (PtBEBT). Both enzymes were also capable of synthesizing plant volatile alcohol esters, such as hexenyl benzoate, at trace levels. Although the activities demonstrated are consistent with roles in salicinoid biosynthesis, direct tests of this hypothesis using transgenic poplar must still be performed.

Topics: Acyl Coenzyme A; Acyltransferases; Benzoates; Benzyl Alcohols; DNA, Complementary; Escherichia coli; Glucosides; Glycosides; Molecular Structure; Phenols; Plant Leaves; Populus

Microbial symbionts are becoming increasingly recognized as mediators of many aspects of plant - herbivore interactions. However, the influence of plant chemical defenses on gut associates of insect herbivores is less well understood. We used gypsy moth (Lymantria dispar L.), and differing trembling aspen (Populus tremuloides Michx.) genotypes that vary in chemical defenses, to assess the influence of foliar chemistry on bacterial communities of larval midguts. We evaluated the bacterial community composition of foliage, and of midguts of larvae feeding on those leaves, using next-generation high-throughput sequencing. Plant defense chemicals did not influence the composition of foliar communities. In contrast, both phenolic glycosides and condensed tannins affected the bacterial consortia of gypsy moth midguts. The two most abundant operational taxonomic units were classified as Ralstonia and Acinetobacter. The relative abundance of Ralstonia was higher in midguts than in foliage when phenolic glycoside concentrations were low, but lower in midguts when phenolic glycosides were high. In contrast, the relative abundance of Ralstonia was lower in midguts than in foliage when condensed tannin concentrations were low, but higher in midguts when condensed tannins were high. Acinetobacter showed a different relationship with host chemistry, being relatively more abundant in midguts than with foliage when condensed tannin concentrations were low, but lower in midguts when condensed tannins were high. Acinetobacter tended to have a greater relative abundance in midguts of insects feeding on genotypes with high phenolic glycoside concentrations. These results show that plant defense chemicals influence herbivore midgut communities, which may in turn influence host utilization.

Topics: Acinetobacter; Animals; Digestive System; Genotype; Glucosides; Herbivory; Larva; Moths; Phenols; Plant Leaves; Populus; Ralstonia; RNA, Ribosomal, 16S; Tannins

We studied the effect of epidermal leaf mining on the leaf chemistry of quaking aspen, Populus tremuloides, during an outbreak of the aspen leaf miner, Phyllocnistis populiella, in the boreal forest of interior Alaska. Phyllocnistis populiella feeds on the epidermal cells of P. tremuloides leaves. Eleven days after the onset of leaf mining, concentrations of the phenolic glycosides tremulacin and salicortin were significantly higher in aspen leaves that had received natural levels of leaf mining than in leaves sprayed with insecticide to reduce mining damage. In a second experiment, we examined the time course of induction in more detail. The levels of foliar phenolic glycosides in naturally mined ramets increased relative to the levels in insecticide-treated ramets on the ninth day following the onset of leaf mining. Induction occurred while some leaf miner larvae were still feeding and when leaves had sustained mining over 5% of the leaf surface. Leaves with extrafloral nectaries (EFNs) had significantly higher constitutive and induced levels of phenolic glycosides than leaves lacking EFNs, but there was no difference in the ability of leaves with and without EFNs to induce phenolic glycosides in response to mining. Previous work showed that the extent of leaf mining damage was negatively related to the total foliar phenolic glycoside concentration, suggesting that phenolic glycosides deter or reduce mining damage. The results presented here demonstrate that induction of phenolic glycosides can be triggered by relatively small amounts of mining damage confined to the epidermal tissue, and that these changes in leaf chemistry occur while a subset of leaf miners are still feeding within the leaf.

Topics: Animals; Glucosides; Host-Parasite Interactions; Moths; Plant Epidermis; Plant Leaves; Plant Nectar; Populus

Understanding the links among plant genotype, plant chemistry, and food selection by vertebrate herbivores is critical to assess the role of herbivores in the evolution of plant secondary chemistry. Some specialized vertebrate herbivores have been shown to select plants differentially according to plant genotype, but examples from generalists, which constitute the vast majority of vertebrate herbivores, are few, especially in natural conditions. We examined the relationship between the North American porcupine (Erethizon dorsatum), a generalist mammalian herbivore, and clonal trembling aspen (Populus tremuloides), a preferred food source of porcupines. We determined preference for certain aspen trees through visual examination of porcupine climbing scars left on tree bark, and through a controlled feeding experiment. We used genetic and biochemical analyses to link the behavioral archives (climbing scars) left by porcupines on aspen trunks to the clonal structure and chemical composition of trees. We show that two phenolic glycosides (tremulacin and salicortin), which are under a high degree of genetic control and thus vary in concentration across clones, are the chemical variables that most influence (deter) feeding choices by porcupines. Using behavioral archives left by a wild herbivore on a natural stand of plants thus allowed us to demonstrate that a generalist vertebrate herbivore can choose plants according to their clonal structure and genetically based chemical composition. Our results contribute to extending previous findings obtained with generalist herbivores studied in controlled conditions, and with specialist herbivores studied in the field.

Topics: Analysis of Variance; Animals; Appetitive Behavior; Feeding Behavior; Glucosides; Plant Bark; Plant Leaves; Populus; Porcupines; Quebec

We have developed a short and practical first synthesis of methyl 1-hydroxy-6-oxo-2-cyclohexenecarboxylate (2), which has been known as a component of salicortin and tremulacin since 1970. Birch reduction of the SEM ether of methyl salicylate followed by oxidation of the intermediate enolate with (-)-camphorsulfonyloxaziridine afforded the SEM enol ether of 2. Hydrolysis of the SEM enol ether afforded 2. We did not observe the dimerization of either racemic or optically enriched 2 to give idesolide (1).

Topics: Carboxylic Acids; Crystallography, X-Ray; Cyclohexenes; Glucosides; Hydrolysis; Models, Molecular; Molecular Structure; Spiro Compounds; Stereoisomerism

Laccases are encoded by multigene families in plants. Previously, we reported the cloning and characterization of five divergent laccase genes from poplar (Populus trichocarpa) xylem. To investigate the role of individual laccase genes in plant development, and more particularly in lignification, three independent populations of antisense poplar plants, lac3AS, lac90AS, and lac110AS with significantly reduced levels of laccase expression were generated. A repression of laccase gene expression had no effect on overall growth and development. Moreover, neither lignin content nor composition was significantly altered as a result of laccase suppression. However, one of the transgenic populations, lac3AS, exhibited a 2- to 3-fold increase in total soluble phenolic content. As indicated by toluidine blue staining, these phenolics preferentially accumulate in xylem ray parenchyma cells. In addition, light and electron microscopic observations of lac3AS stems indicated that lac3 gene suppression led to a dramatic alteration of xylem fiber cell walls. Individual fiber cells were severely deformed, exhibiting modifications in fluorescence emission at the primary wall/middle lamella region and frequent sites of cell wall detachment. Although a direct correlation between laccase gene expression and lignification could not be assigned, we show that the gene product of lac3 is essential for normal cell wall structure and integrity in xylem fibers. lac3AS plants provide a unique opportunity to explore laccase function in plants.

Topics: Benzyl Alcohols; Cell Wall; DNA, Antisense; Down-Regulation; Gene Expression Regulation, Plant; Glucosides; Laccase; Light; Lignin; Microscopy, Electron; Molecular Structure; Multigene Family; Oxidoreductases; Phenols; Plants, Genetically Modified; RNA, Messenger; Salicaceae; Spectrum Analysis