02821nas a2200325   4500008004100000020001400041245014800055210006900203260000800272300001200280490000700292520173200299653001502031100002902046700002202075700001402097700001502111700002002126700002302146700002502169700002002194700002202214700001702236700002002253700002102273700002102294700002502315700002002340856013502360       2012                        eng d  a0965-174800aTranscriptome and full-length cDNA resources for the mountain pine beetle, Dendroctonus ponderosae Hopkins, a major insect pest of pine forests0 aTranscriptome and fulllength cDNA resources for the mountain pin  cAug  a525-5360 v423 aBark beetles (Coleoptera: Curculionidae: Scolytinae) are major insect pests of many woody plants around the world. The mountain pine beetle (MPB), Dendroctonus ponderosae Hopkins, is a significant historical pest of western North American pine forests. It is currently devastating pine forests in western North America - particularly in British Columbia, Canada - and is beginning to expand its host range eastward into the Canadian boreal forest, which extends to the Atlantic coast of North America. Limited genomic resources are available for this and other bark beetle pests, restricting the use of genomics-based information to help monitor, predict, and manage the spread of these insects. To overcome these limitations, we generated comprehensive transcriptome resources from fourteen full-length enriched cDNA libraries through paired-end Sanger sequencing of 100,000 cDNA clones, and single-end Roche 454 pyrosequencing of three of these cDNA libraries. Hybrid de novo assembly of the 3.4 million sequences resulted in 20,571 isotigs in 14,410 isogroups and 246,848 singletons. In addition, over 2300 non-redundant full-length cDNA clones putatively containing complete open reading frames, including 47 cytochrome P450s, were sequenced fully to high quality. This first large-scale genomics resource for bark beetles provides the relevant sequence information for gene discovery; functional and population genomics; comparative analyses; and for future efforts to annotate the MPB genome. These resources permit the study of this beetle at the molecular level and will inform research in other Dendroctonus spp. and more generally in the Curculionidae and other Coleoptera. (C) 2012 Elsevier Ltd. All rights reserved.10aScolytinae1 aKeeling, Christopher, I.1 aHenderson, Hannah1 aLi, Maria1 aYuen, Mack1 aClark, Erin, L.1 aFraser, Jordie, D.1 aHuber, Dezene, P. W.1 aLiao, Nancy, Y.1 aDocking, Roderick1 aBirol, Inanc1 aChan, Simon, K.1 aTaylor, Greg, A.1 aPalmquist, Diana1 aJones, Steven, J. M.1 aBohlmann, Joerg  uhttps://weevil.myspecies.info/transcriptome-and-full-length-cdna-resources-mountain-pine-beetle-dendroctonus-ponderosae-hopkins-ma02905nas a2200205   4500008004100000020001400041245014700055210006900202260000800271300001000279490000700289520211800296100002902414700002502443700002502468700002202493700002202515700002702537856013502564       2012                        eng d  a0965-174800aRecombinant expression, localization and in vitro inhibition of midgut cysteine peptidase (Sl-CathL) from sugarcane weevil, Sphenophorus levis0 aRecombinant expression localization and in vitro inhibition of m  cJan  a58-690 v423 aA cDNA coding for a digestive cathepsin L, denominated Sl-CathL, was isolated from a cDNA library of Sphenophorus levis larvae, representing the most abundant EST (10.49%) responsible for proteolysis in the midgut. The open reading frame of 972 bp encodes a preproenzyme similar to midgut cathepsin L-like enzymes in other coleopterans. Recombinant Sl-CathL was expressed in Pichia pastoris, with molecular mass of about 42 kDa. The recombinant protein was catalytically activated at low pH and the mature enzyme of 39 kDa displayed thermal instability and maximal activity at 37 [degree]C and pH 6.0. Immunocytochemical analysis revealed Sl-CathL production in the midgut epithelium and secretion from vesicles containing the enzyme into the gut lumen, confirming an important role for this enzyme in the digestion of the insect larvae. The expression profile identified by RT-PCR through the biological cycle indicates that Sl-CathL is mainly produced in larval stages, with peak expression in 30-day-old larvae. At this stage, the enzyme is 1250-fold more expressed than in the pupal fase, in which the lowest expression level is detected. This enzyme is also produced in the adult stage, albeit in lesser abundance, assuming the presence of a different array of enzymes in the digestive system of adults. Tissue-specific analysis revealed that Sl-CathL mRNA synthesis occurs fundamentally in the larval midgut, thereby confirming its function as a digestive enzyme, as detected in immunolocalization assays. The catalytic efficiency of the purified recombinant enzyme was calculated using different substrates (Z-Leu-Arg-AMC, Z-Arg-Arg-AMC and Z-Phe-Arg-AMC) and rSl-CathL exhibited hydrolysis preference for Z-Leu-Arg-AMC (kcat/Km = 37.53 mM S-1), which is similar to other insect cathepsin L-like enzymes. rSl-CathL activity inhibition assays were performed using four recombinant sugarcane cystatins. rSl-CathL was strongly inhibited by recombinant cystatin CaneCPI-4 (Ki = 0.196 nM), indicating that this protease is a potential target for pest control. [copyright] 2011 Elsevier Ltd. All rights reserved.1 aFonseca, Fernando, P. P.1 aSoares-Costa, Andrea1 aRibeiro, Alberto, F.1 aRosa, Jose, Cesar1 aTerra, Walter, R.1 aHenrique-Silva, Flavio  uhttps://weevil.myspecies.info/recombinant-expression-localization-and-vitro-inhibition-midgut-cysteine-peptidase-sl-cathl-sugarcan02118nas a2200181   4500008004100000020001400041245010500055210006900160260000800229300001000237490000700247520146400254100002501718700002301743700002401766700002001790856012601810       2012                        eng d  a0965-174800aIpsdienol dehydrogenase (IDOLDH): A novel oxidoreductase important for Ips pini pheromone production0 aIpsdienol dehydrogenase IDOLDH A novel oxidoreductase important   cFeb  a81-900 v423 aIpsdienone (2-methyl-6-methylene-2,7-octadien-4-one) is an important intermediate in the biosynthesis of pheromonal ipsdienol (2-methyl-6-methylene-2,7-octadien-4-ol) and ipsenol (2-methyl-6methylene-7-octen-4-ol) in male pine engraver beetles. Ips pini (Say). A novel ipsdienol dehydrogenase (IDOLDH) with a pheromone-biosynthetic gene expression pattern was cloned, expressed, functionally characterized, and its cellular localization analyzed. The cDNA has a 762 nt ORF encoding a 253 amino acid predicted translation product of 28 kDa and pI 5.8. The protein has conserved motifs of the Cp2 subfamily of "classical" short-chain dehydrogenases. Transcript levels were highest in pheromone producing tissue: the anterior midgut of fed males. The protein was detected only in male midguts and localized in the cytosolic fraction of midgut cells. Recombinant IDOLDH was produced in Sf9 cells using a baculovirus expression system. Enzyme assays of protein preparations showed IDOLDH used both NAD+ and NADP+ as coenzymes with specific activities in the nanomole range. Enzyme assays and GC/MS analysis showed that IDOLDH catalyzed the oxidation of racemic ipsdienol and (4R)-(-)-ipsdienol to form ipsdienone, while (4S)-(+)-ipsdienol was not a substrate. These data strongly implicate IDOLDH as an enzyme involved in terminal pheromone-biosynthetic steps, likely functioning to "tune" ipsdienol enantiomeric ratios. (C) 2011 Elsevier Ltd. All rights reserved.1 aFigueroa-Teran, Rubi1 aWelch, William, H.1 aBlomquist, Gary, J.1 aTirtiger, Claus  uhttps://weevil.myspecies.info/ipsdienol-dehydrogenase-idoldh-novel-oxidoreductase-important-ips-pini-pheromone-production02329nas a2200169   4500008004100000020001400041245014800055210006900203260000800272300001200280490000700292520165600299100002001955700002401975700002501999856013502024       2010                        eng d  a0965-174800aMolecular and biochemical characterisation of a dual proteolytic system in vine weevil larvae (Otiorhynchus sulcatus Coleoptera: Curculionidae)0 aMolecular and biochemical characterisation of a dual proteolytic  cNov  a785-7910 v403 aThe ability of phytophagous insects to utilise the relatively low nitrogen content of plant tissues is typically the limiting factor in their nutritional uptake. In the larval stage, the vine weevil feeds predominantly on root tissues of plants. The root tissue as a whole has low levels of free amino acids, and thus effective hydrolysis of dietary proteins is essential for survival. In contrast to previous reports the present study demonstrates through both molecular and biochemical studies the presence of proteolytic enzymes from two mechanistic classes, cysteine and serine proteases, in the gut of larval vine weevil; with the latter being the predominant form. cDNA clones encoding cathepsin B-like and serine-like sequences were isolated from a gut specific cDNA library; the cathepsin B-like clone has the Cys-His-Asn catalytic triad. However, the sequence showed the replacement of the conserved His His sequence in the "occluding loop" region of the enzyme with Asp His. This may result in a change to the substrate specificity. Two trypsin precursors contained evidence of a signal peptide, activation peptide, and conserved N-termini (IVGG). Other structural features included typical His, Asp. and Set residues of the catalytic amino acid triad indicative of serine proteases, characteristic residues in the substrate-binding pocket, and four pairs of cysteine residues for disulfide bridges. The apparent abundance of the trypsin-like cDNA clones compared to the cathepsin B clones suggests that serine proteases are the predominant form, thus supporting data from the biochemical studies. (C) 2010 Elsevier Ltd. All rights reserved.1 aEdwards, M., G.1 aGatehouse, John, A.1 aGatehouse, A., M. R.  uhttps://weevil.myspecies.info/molecular-and-biochemical-characterisation-dual-proteolytic-system-vine-weevil-larvae-otiorhynchus-s03689nas a2200241   4500008004100000020001400041245004100055210004100096260000800137300001200145490000700157520304700164653000803211100002403219700002503243700001303268700001703281700002203298700002303320700001603343700002003359856006803379       2010                        eng d  a0965-174800aPheromone production in bark beetles0 aPheromone production in bark beetles  cOct  a699-7120 v403 aThe first aggregation Pheromone components from bark beetles were identified in 1966 as a mixture of ipsdienol, ipsenol and verbenol. Since then, a number of additional components have been identified as both aggregation and anti-aggregation pheromones, with many of them being monoterpenoids or derived from monoterpenoids. The structural similarity between the major pheromone components of bark beetles and the monoterpenes found in the host trees, along with the association of monoterpenoid production with plant tissue, led to the paradigm that most if not all bark beetle pheromone components were derived from host tree precursors, often with a simple hydroxylation producing the pheromone. In the 1990s there was a paradigm shift as evidence for de novo biosynthesis of pheromone components began to accumulate, and it is now recognized that most bark beetle monoterpenoid aggregation pheromone components are biosynthesized de novo. The bark beetle aggregation pheromones are released from the frass, which is consistent with the isoprenoid aggregation pheromones, including ipsdienol, ipsenol and frontalin, being produced in midgut tissue. It appears that exo-brevocomin is produced de novo in fat body tissue, and that verbenol, verbenone and verbenene are produced from dietary cc-pinene in fat body tissue. Combined biochemical, molecular and functional genomics studies in Ips pini yielded the discovery and characterization of the enzymes that convert mevalonate pathway intermediates to pheromone components, including a novel bifunctional geranyl diphosphate synthase/myrcene synthase, a cytochrome P450 that hydroxylates myrcene to ipsdienol, and an oxidoreductase that interconverts ipsdienol and ipsdienone to achieve the appropriate stereochemistry of ipsdienol for pheromonal activity. Furthermore, the regulation of these genes and their corresponding enzymes proved complex and diverse in different species. Mevalonate pathway genes in pheromone producing male I. pini have much higher basal levels than in females, and feeding induces their expression. In I. dupIicatus and I. pini, juvenile hormone III (JH III) induces pheromone production in the absence of feeding, whereas in I. paraconfusus and I. confusus, topically applied JH III does not induce pheromone production. In all four species, feeding induces pheromone production. While many of the details of pheromone production, including the site of synthesis, pathways and knowledge of the enzymes involved are known for Ips, less is known about pheromone production in Dendroctonus. Functional genomics studies are under way in D. ponderosae, which should rapidly increase our understanding of pheromone production in this genus. This chapter presents a historical development of what is known about pheromone production in bark beetles, emphasizes the genomic and post-genomic work in I. pini and points out areas where research is needed to obtain a more complete understanding of pheromone production. [copyright]2010 Elsevier Ltd. All rights reserved.10aIps1 aBlomquist, Gary, J.1 aFigueroa-Teran, Rubi1 aAw, Mory1 aSong, Minmin1 aGorzalski, Andrew1 aAbbott, Nicole, L.1 aChang, Eric1 aTittiger, Claus  uhttps://weevil.myspecies.info/pheromone-production-bark-beetles02128nas a2200217   4500008004500000020001400045245016300059210006900222260001300291300001200304490000700316520132900323653001301652100002201665700001601687700002001703700001401723700002201737700001701759856013401776       2008                        Engldsh    a0965-174800aIsolation, endocrine regulation and transcript distribution of a putative primary JH-responsive gene from the pine engraver, Ips pini (Coleoptera: Scolytidae)0 aIsolation endocrine regulation and transcript distribution of a   cFebruary  a256-2670 v383 aWe isolated a cDNA of unknown function from a juvenile hormone III (JH III)-treated male midgut cDNA library prepared from the pine engraver beetle, Ips pini, and examined its genomic structure. The gene, tentatively named "Ipi10G08", encoded a 4 10 amino acid translation product that shared 26-37% identity with unannotated matches from several insects. Semi-quantitative RT-PCR analysis of Ipi10G08 following application of a 10 [mu]g dose of JH III demonstrated an early induction for both male and female beetles, with transcripts being detectable after 45 min. An expression profile of male midgut tissue indicated Ipi10G08 transcript levels reach a maximum induction of [approximately]22.5-fold control levels at 4 h post-treatment. Tissue distribution studies displayed a large induction of Ipi10G08 mRNA in the alimentary canal of JH Ill-treated beetles, especially in males. A dose curve from both sexes suggested there may be a difference in the ability to respond to lower levels of JH III and immunoblot analysis indicated that although JH III highly induces transcript levels in females, protein levels are not similarly induced, while protein levels are induced in males. Ipi10G08 is likely a primary JH response gene and may provide insight into how this hormone exerts its actions. Published by Elsevier Ltd10aIps pini1 aBearfield, J., C.1 aBox, C., D.1 aKeeling, C., I.1 aYoung, S.1 aBlomquist, G., J.1 aTittiger, C.  uhttps://weevil.myspecies.info/isolation-endocrine-regulation-and-transcript-distribution-putative-primary-jh-responsive-gene-pine01962nas a2200265   4500008004100000020001400041245018800055210006900243260000800312300001200320490000700332520093200339653002301271100002701294700002601321700002801347700002701375700003001402700003201432700002701464700001901491700002101510700003001531856013501561       2010                        eng d  a0965-174800aMidgut GPI-anchored proteins with alkaline phosphatase activity from the cotton boll weevil (Anthonomus grandis) are putative receptors for the Cry1B protein of Bacillus thuringiensis0 aMidgut GPIanchored proteins with alkaline phosphatase activity f  cFeb  a138-1450 v403 aCry toxins from Bacillus thuringiensis (Bt) are used for insect control. They interact with specific receptors located on the host cell surface and are activated by host proteases following receptor binding resulting in midgut epithelial cells lysis. In this work we had cloned, sequenced and expressed a cry1Ba toxin gene from the B thuringiensis S601 strain which was previously shown to be toxic to Anthonomus grandis, a cotton pest. The Cry1Ba6 protein expressed in an acrystaliferous B. thuringiensis strain was toxic to A. grandis in bioassays. The binding of Cry1Ba6 toxin to proteins located in the midgut brush border membrane of A. grandis was analyzed and we found that Cry1Ba6 binds to two proteins (62 and 65 kDa) that showed alkaline phosphatase (ALP) activity. This work is the first report that shows the localization of Cry toxin receptors in the midgut cells of A. grandis. (C) 2009 Published by Elsevier Ltd.10aAnthonomus grandis1 aMartins, Erica, Soares1 aMonnerat, Rose, Gomes1 aQueiroz, Paulo, Roberto1 aDumas, Vinicius, Fiuza1 aBraz, Shelida, Vasconcelo1 aAguiar, Raimundo, Wagner de1 aGomes, Ana, Cristina M1 aSanchez, Jorge1 aBravo, Alejandra1 aRibeiro, Bergmann, Morais  uhttps://weevil.myspecies.info/midgut-gpi-anchored-proteins-alkaline-phosphatase-activity-cotton-boll-weevil-anthonomus-grandis-are