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  Subjects -> BIOLOGY (Total: 2698 journals)
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BIOCHEMISTRY (206 journals)                  1 2 3     

AAPS PharmSciTech     Hybrid Journal   (Followers: 7)
Acetic Acid Bacteria     Open Access   (Followers: 1)
ACS Chemical Biology     Full-text available via subscription   (Followers: 342)
ACS Chemical Neuroscience     Full-text available via subscription   (Followers: 13)
Acta Crystallographica Section D : Biological Crystallography     Hybrid Journal   (Followers: 9)
Acta Crystallographica Section F: Structural Biology Communications     Hybrid Journal   (Followers: 6)
Advances and Applications in Bioinformatics and Chemistry     Open Access   (Followers: 8)
Advances in Biological Chemistry     Open Access   (Followers: 5)
Advances in Carbohydrate Chemistry and Biochemistry     Full-text available via subscription   (Followers: 8)
Advances in Plant Biochemistry and Molecular Biology     Full-text available via subscription   (Followers: 7)
Advances in Protein Chemistry and Structural Biology     Full-text available via subscription   (Followers: 10)
African Journal of Biochemistry Research     Open Access  
African Journal of Chemical Education     Open Access   (Followers: 1)
Alkaloids: Chemical and Biological Perspectives     Full-text available via subscription   (Followers: 4)
American Journal of Biochemistry     Open Access   (Followers: 6)
American Journal of Biochemistry and Biotechnology     Open Access   (Followers: 201)
American Journal of Biochemistry and Molecular Biology     Open Access   (Followers: 11)
American Journal of Polymer Science     Open Access   (Followers: 17)
Amino Acids     Hybrid Journal   (Followers: 7)
Analytical Biochemistry     Hybrid Journal   (Followers: 231)
Annals of Clinical Biochemistry     Hybrid Journal   (Followers: 1)
Annual Review of Biochemistry     Full-text available via subscription   (Followers: 29)
Annual Review of Chemical and Biomolecular Engineering     Full-text available via subscription   (Followers: 10)
Applied Biochemistry and Biotechnology     Hybrid Journal   (Followers: 17)
Applied Biochemistry and Microbiology     Hybrid Journal   (Followers: 7)
Applied Organometallic Chemistry     Hybrid Journal   (Followers: 4)
Archives of Biochemistry and Biophysics     Hybrid Journal   (Followers: 9)
Archives of Insect Biochemistry and Physiology     Hybrid Journal   (Followers: 1)
Archives Of Physiology And Biochemistry     Hybrid Journal   (Followers: 1)
Asian Journal of Biochemistry     Open Access   (Followers: 1)
Asian Journal of Biomedical and Pharmaceutical Sciences     Open Access   (Followers: 1)
Avicenna Journal of Medical Biochemistry     Open Access  
Bangladesh Journal of Medical Biochemistry     Open Access   (Followers: 2)
BBA Clinical     Open Access  
BBR : Biochemistry and Biotechnology Reports     Open Access   (Followers: 3)
Biocatalysis     Open Access  
Biochemical and Biophysical Research Communications     Hybrid Journal   (Followers: 15)
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Biochemical Compounds     Open Access  
Biochemical Engineering Journal     Hybrid Journal   (Followers: 9)
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Biochemical Journal     Full-text available via subscription   (Followers: 16)
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Biochemical Society Transactions     Full-text available via subscription   (Followers: 3)
Biochemical Systematics and Ecology     Hybrid Journal   (Followers: 3)
Biochemistry     Full-text available via subscription   (Followers: 253)
Biochemistry (Moscow)     Hybrid Journal   (Followers: 3)
Biochemistry (Moscow) Supplement Series A: Membrane and Cell Biology     Hybrid Journal   (Followers: 4)
Biochemistry (Moscow) Supplemental Series B: Biomedical Chemistry     Hybrid Journal   (Followers: 3)
Biochemistry and Cell Biology     Full-text available via subscription   (Followers: 8)
Biochemistry and Molecular Biology Education     Hybrid Journal   (Followers: 3)
Biochemistry and Molecular Biology of Fishes     Full-text available via subscription   (Followers: 1)
Biochemistry Research International     Open Access   (Followers: 4)
Biochimica et Biophysica Acta (BBA) - Molecular and Cell Biology of Lipids     Hybrid Journal   (Followers: 3)
Biochimica et Biophysica Acta (BBA) - Molecular Basis of Disease     Hybrid Journal   (Followers: 18)
Biochimica et Biophysica Acta (BBA) - Molecular Cell Research     Hybrid Journal   (Followers: 6)
Biochimie     Hybrid Journal   (Followers: 4)
Bioconjugate Chemistry     Full-text available via subscription   (Followers: 14)
BioDrugs     Full-text available via subscription   (Followers: 7)
Bioelectrochemistry     Hybrid Journal   (Followers: 3)
Biofuels     Hybrid Journal   (Followers: 8)
Biogeochemistry     Hybrid Journal   (Followers: 7)
BioInorganic Reaction Mechanisms     Full-text available via subscription   (Followers: 1)
Biokemistri     Open Access  
Biological Chemistry     Partially Free   (Followers: 11)
Biomaterials Research     Open Access  
Biomedicines     Open Access   (Followers: 1)
BioMolecular Concepts     Full-text available via subscription   (Followers: 2)
Bioprocess     Open Access  
Bioscience, Biotechnology, and Biochemistry     Hybrid Journal   (Followers: 6)
Biosimilars     Open Access   (Followers: 1)
Biotechnology and Applied Biochemistry     Hybrid Journal   (Followers: 18)
BMC Biochemistry     Open Access   (Followers: 8)
BMC Chemical Biology     Open Access   (Followers: 4)
Bulletin of University of Agricultural Sciences and Veterinary Medicine Cluj-Napoca : Food Science and Technology     Open Access  
Carbohydrate Polymers     Hybrid Journal   (Followers: 9)
Cell Biochemistry and Biophysics     Hybrid Journal   (Followers: 6)
Cell Biochemistry and Function     Hybrid Journal   (Followers: 3)
Cellular Physiology and Biochemistry     Open Access   (Followers: 3)
Central European Journal of Chemistry     Hybrid Journal   (Followers: 5)
ChemBioChem     Hybrid Journal   (Followers: 2)
Chemical and Biological Technologies for Agriculture     Open Access  
Chemical Biology & Drug Design     Hybrid Journal   (Followers: 23)
Chemical Engineering Journal     Hybrid Journal   (Followers: 20)
Chemical Senses     Hybrid Journal   (Followers: 1)
Chemical Speciation and Bioavailability     Full-text available via subscription   (Followers: 1)
Chemico-Biological Interactions     Hybrid Journal   (Followers: 2)
Chemistry & Biodiversity     Hybrid Journal   (Followers: 5)
Chemistry & Biology     Full-text available via subscription   (Followers: 17)
Chemistry and Ecology     Hybrid Journal   (Followers: 1)
Clinical Biochemist Reviews     Full-text available via subscription   (Followers: 1)
Clinical Biochemistry     Hybrid Journal   (Followers: 3)
Clinical Chemistry and Laboratory Medicine     Full-text available via subscription   (Followers: 6)
Clinical Lipidology     Full-text available via subscription  
Comparative Biochemistry and Physiology Part A: Molecular & Integrative Physiology     Hybrid Journal   (Followers: 5)
Comparative Biochemistry and Physiology Part B: Biochemistry and Molecular Biology     Hybrid Journal   (Followers: 2)
Comparative Biochemistry and Physiology Part C: Toxicology & Pharmacology     Hybrid Journal   (Followers: 5)
Comparative Biochemistry and Physiology Part D: Genomics and Proteomics     Hybrid Journal   (Followers: 3)
Comprehensive Biochemistry     Full-text available via subscription   (Followers: 1)
Computational Biology and Chemistry     Hybrid Journal   (Followers: 8)

        1 2 3     

Journal Cover Insect Biochemistry and Molecular Biology     [SJR: 1.333]   [H-I: 69]
   [5 followers]  Follow    
   Hybrid Journal Hybrid journal (It can contain Open Access articles)
   ISSN (Print) 0965-1748
   Published by Elsevier Homepage  [2585 journals]
  • Overview of chitin metabolism enzymes in Manduca sexta: Identification,
           domain organization, phylogenetic analysis and gene expression
    • Abstract: Publication date: Available online 20 January 2015
      Source:Insect Biochemistry and Molecular Biology
      Author(s): Guillaume Tetreau , Xiaolong Cao , Yun-Ru Chen , Subbaratnam Muthukrishnan , Jiang Haobo , Gary W. Blissard , Michael R. Kanost , Ping Wang
      Chitin is one of the most abundant biomaterials in nature. The biosynthesis and degradation of chitin in insects are complex and dynamically regulated to cope with insect growth and development. Chitin metabolism in insects is known to involve numerous enzymes, including chitin synthases (synthesis of chitin), chitin deacetylases (modification of chitin by deacetylation) and chitinases (degradation of chitin by hydrolysis). In this study, we conducted a genome-wide search and analysis of genes encoding these chitin metabolism enzymes in Manduca sexta. Our analysis confirmed that only two chitin synthases are present in M. sexta as in most other arthropods. Eleven chitin deacetylases (encoded by nine genes) were identified, with at least one representative in each of the five phylogenetic groups that have been described for chitin deacetylases to date. Eleven genes encoding for family 18 chitinases (GH18) were found in the M. sexta genome. Based on the presence of conserved sequence motifs in the catalytic sequences and phylogenetic relationships, two of the M. sexta chitinases did not cluster with any of the current eight phylogenetic groups of chitinases: two new groups were created (groups IX and X) and their characteristics are described. The result of the analysis of the Lepidoptera-specific chitinase-h (group h) is consistent with its proposed bacterial origin. By analyzing chitinases from fourteen species that belong to seven different phylogenetic groups, we reveal that the chitinase genes appear to have evolved sequentially in the arthropod lineage to achieve the current high level of diversity observed in M. sexta. Based on the sequence conservation of the catalytic domains and on their developmental stage- and tissue-specific expression, we propose putative functions for each group in each category of enzymes.
      Graphical abstract image

      PubDate: 2015-01-24T08:30:34Z
       
  • Integrated modeling of protein-coding genes in the Manduca sexta genome
           using RNA-Seq data from the biochemical model insect
    • Abstract: Publication date: Available online 20 January 2015
      Source:Insect Biochemistry and Molecular Biology
      Author(s): Xiaolong Cao , Haobo Jiang
      The genome sequence of Manduca sexta was recently determined using 454 technology. Cufflinks and MAKER2 were used to establish gene models in the genome assembly based on the RNA-Seq data and other species' sequences. Aided by the extensive RNA-Seq data from 50 tissue samples at various life stages, annotators over the world (including the present authors) have manually confirmed and improved a small percentage of the models after spending months of effort. While such collaborative efforts are highly commendable, many of the predicted genes still have problems which may hamper future research on this insect species. As a biochemical model representing lepidopteran pests, M. sexta has been used extensively to study insect physiological processes for over five decades. In this work, we assembled Manduca datasets Cufflinks 3.0, Trinity 4.0, and Oases 4.0 to assist the manual annotation efforts and development of Official Gene Set (OGS) 2.0. To further improve annotation quality, we developed methods to evaluate gene models in the MAKER2, Cufflinks, Oases and Trinity assemblies and selected the best ones to constitute MCOT 1.0 after thorough crosschecking. MCOT 1.0 has 18,089 genes encoding 31,666 proteins: 32.8% match OGS 2.0 models perfectly or near perfectly, 11,747 differ considerably, and 29.5% are absent in OGS 2.0. Future automation of this process is anticipated to greatly reduce human efforts in generating comprehensive, reliable models of structural genes in other genome projects where extensive RNA-Seq data are available.
      Graphical abstract image

      PubDate: 2015-01-24T08:30:34Z
       
  • Two chitinase 5 genes from Locusta migratoria: molecular characteristics
           and functional differentiation
    • Abstract: Publication date: Available online 23 January 2015
      Source:Insect Biochemistry and Molecular Biology
      Author(s): Daqi Li , Jianqin Zhang , Yan Wang , Xiaojian Liu , Enbo Ma , Yi Suna , Sheng Li , Kun Yan Zhu , Jianzhen Zhang
      The duplication of chitinase 5 (Cht5) into two to five different genes has been reported only in mosquito species to date. Here, we report the duplication of Cht5 genes (LmCht5-1 and LmCht5-2) in the migratory locust (Locusta migratoria). Both LmCht5-1 (505 aa) and LmCht5-2 (492 aa) possess a signal peptide and a catalytic domain with four conserved motifs, but only LmCht5-1 contains a chitin-binding domain. Structural and phylogenetic analyses suggest that LmCht5-1 is orthologous to other insect Cht5 genes, whereas LmCht5-2 might be newly duplicated. Both LmCht5 genes were expressed in all tested tissues with LmCht5-1 highly expressed in hindgut and LmCht5-2 highly expressed in integument, foregut, hindgut and fat bodies. From the fourth-instar nymphs to the adults, LmCht5-1 and LmCht5-2 showed similar developmental expression patterns with transcript peaks prior to each nymphal molting, suggesting that their expression levels are similarly regulated. Treatment with 20-hydroxyecdysone (20E; the most active molting hormone) and reducing expression of EcR (ecdysone receptor gene) by RNAi increased and decreased expression of both LmCht5 genes, respectively, indicating that both genes are responsive to 20E. Although transcript level of LmCht5-2 is generally 10-fold higher than that of LmCht5-1, RNAi-mediated suppression of LmCht5-1 transcript led to severe molting defects and lethality, but such effects were not seen with RNAi of LmCht5-2, suggesting that the newly duplicated LmCht5-2 is not essential for development and survivorship of the locust.
      Graphical abstract image

      PubDate: 2015-01-24T08:30:34Z
       
  • Biochemical characterization of maintenance DNA methyltransferase DNMT-1
           from silkworm, Bombyx mori
    • Abstract: Publication date: Available online 23 January 2015
      Source:Insect Biochemistry and Molecular Biology
      Author(s): Takumi Mitsudome , Hiroaki Mon , Jian Xu , Zhiqing Li , Jae Man Lee , Anandrao Ashok Patil , Atsushi Masuda , Kazuhiro Iiyama , Daisuke Morokuma , Takahiro Kusakabe
      DNA methylation is an important epigenetic mechanism involved in gene expression of vertebrates and invertebrates. In general, DNA methylation profile is established by de novo DNA methyltransferases (DNMT-3A, -3B) and maintainance DNA methyltransferase (DNMT-1). DNMT-1 has a strong substrate preference for hemimethylated DNA over the unmethylated one. Because the silkworm genome lacks an apparent homologue of de novo DNMT, it is still unclear that how silkworm chromosome establishes and maintains its DNA methylation profile. As the first step to unravel this enigma, we purified recombinant BmDNMT-1 using baculovirus expression system and characterized its DNA-binding and DNA methylation activity. We found that the BmDNMT-1 preferentially methylates hemimethylated DNA despite binding to both unmethylated and hemimethylated DNA. Interestingly, BmDNMT-1 formed a complex with DNA in the presence or absence of methyl group donor, S-Adenosylmethionine (AdoMet) and the AdoMet-dependent complex formation was facilitated by Zn2+ and Mn2+. Our results provide clear evidence that BmDNMT-1 retained the function as maintenance DNMT but its sensitivity to metal ions is different from mammalian DNMT-1.
      Graphical abstract image

      PubDate: 2015-01-24T08:30:34Z
       
  • Structural features, evolutionary relationships, and transcriptional
           regulation of C-type lectin-domain proteins in Manduca sexta
    • Abstract: Publication date: Available online 29 December 2014
      Source:Insect Biochemistry and Molecular Biology
      Author(s): Xiang-Jun Rao , Xiaolong Cao , Yan He , Yingxia Hu , Xiufeng Zhang , Yun-Ru Chen , Gary Blissard , Michael R. Kanost , Xiao-Qiang Yu , Haobo Jiang
      C-type lectins (CTLs) are a large family of Ca2+-dependent carbohydrate-binding proteins recognizing various glycoconjugates and functioning primarily in immunity and cell adhesion. We have identified 34 CTLDP (for CTL-domain protein) genes in the Manduca sexta genome, which encode proteins with one to three CTL domains. CTL-S1 through S9 (S for simple) have one or three CTL domains; immulectin-1 through 19 have two CTL domains; CTL-X1 through X6 (X for complex) have one or two CTL domains along with other structural modules. Nine simple CTLs and seventeen immulectins have a signal peptide and are likely extracellular. Five complex CTLs have both an N-terminal signal peptide and a C-terminal transmembrane region, indicating that they are membrane anchored. Immulectins exist broadly in Lepidoptera and lineage-specific gene duplications have generated three clusters of fourteen genes in the M. sexta genome, thirteen of which have similar expression patterns. In contrast to the family expansion, CTL-S1∼S6, S8, and X1∼X6 have 1:1 orthologs in at least four lepidopteran/dipteran/coleopteran species, suggestive of conserved functions in a wide range of holometabolous insects. Structural modeling suggests the key residues for Ca2+-dependent or independent binding of certain carbohydrates by CTL domains. Promoter analysis identified putative κB motifs in eighteen of the CTL genes, which did not have a strong correlation with immune inducibility in the mRNA or protein levels. Together, the gene identification, sequence comparisons, structure modeling, phylogenetic analysis, and expression profiling establish a solid foundation for future studies of M. sexta CTL-domain proteins.
      Graphical abstract image

      PubDate: 2015-01-19T18:28:59Z
       
  • Sequence conservation, phylogenetic relationships, and expression profiles
           of nondigestive serine proteases and serine protease homologs in Manduca
           sexta
    • Abstract: Publication date: Available online 18 December 2014
      Source:Insect Biochemistry and Molecular Biology
      Author(s): Xiaolong Cao , Yan He , Yingxia Hu , Xiufeng Zhang , Yang Wang , Zhen Zou , Yunru Chen , Gary W. Blissard , Michael R. Kanost , Haobo Jiang
      Serine protease (SP) and serine protease homolog (SPH) genes in insects encode a large family of proteins involved in digestion, development, immunity, and other processes. While 68 digestive SPs and their close homologs are reported in a companion paper (Kuwar et al., in preparation), we have identified 125 other SPs/SPHs in Manduca sexta and studied their structure, evolution, and expression. Fifty-two of them contain cystine-stabilized structures for molecular recognition, including clip, LDLa, Sushi, Wonton, TSP, CUB, Frizzle, and SR domains. There are nineteen groups of genes evolved from relatively recent gene duplication and sequence divergence. Thirty-five SPs and seven SPHs contain 1, 2 or 5 clip domains. Multiple sequence alignment and molecular modeling of the 54 clip domains have revealed structural diversity of these regulatory modules. Sequence comparison with their homologs in Drosophila melanogaster, Anopheles gambiae and Tribolium castaneum allows us to classify them into five subfamilies: A are SPHs with 1 or 5 group-3 clip domains, B are SPs with 1 or 2 group-2 clip domains, C, D1 and D2 are SPs with a single clip domain in group-1a, 1b and 1c, respectively. We have classified into six categories the 125 expression profiles of SP-related proteins in fat body, brain, midgut, Malpighian tubule, testis, and ovary at different stages, suggesting that they participate in various physiological processes. Through RNA-Seq-based gene annotation and expression profiling, as well as intragenomic sequence comparisons, we have established a framework of information for future biochemical research of nondigestive SPs and SPHs in this model species.
      Graphical abstract image

      PubDate: 2015-01-19T18:28:59Z
       
  • Spatial and temporal synthesis of Mamestra configurata peritrophic matrix
           through a larval stadium
    • Abstract: Publication date: November 2014
      Source:Insect Biochemistry and Molecular Biology, Volume 54
      Author(s): Umut Toprak , Dwayne D. Hegedus , Doug Baldwin , Cathy Coutu , Martin Erlandson
      The structure and synthesis of the Mamestra configurata peritrophic matrix (PM) was examined at various time points during a larval stadium. Bright field and confocal fluorescence microscopy revealed major differences between the PM of feeding and molting larvae. The PM from feeding larvae was thinner and composed of approximately 5–10 layers. In contrast, mid-molt larvae had a chitinaceaous PM composed of multiple thick layers which filled most of the midgut lumen. PM synthesis initiates in the anterior midgut, based on the expression of genes encoding chitin synthase-2 (CHS-2), coincident with the incorporation of the major structural PM proteins (McIIM1, McIIM2 and McPM1). This is followed by reinforcement with other PM proteins (McIIM3 and McIIM4) as it moves toward the posterior of the midgut. Chitin deacetylase (McCDA1) was associated only with the anterior PM. Collectively, these findings indicate that the structural properties of the PM differ along the length of the midgut. Genes encoding chitinolytic enzymes (McCHI and McNAG) were expressed and exochitinase activity was present when the PM had degraded (pre-molt) and when the new PM was forming (mid-molt), indicating that they are involved in either PM turnover and/or maintenance dependent upon the stage.
      Graphical abstract image

      PubDate: 2015-01-19T18:28:59Z
       
  • Sumoylation modulates 20-hydroxyecdysone signaling by maintaining USP
           protein levels in Drosophila
    • Abstract: Publication date: November 2014
      Source:Insect Biochemistry and Molecular Biology, Volume 54
      Author(s): Jiawan Wang , Sheng Wang , Sheng Li
      The nuclear receptor complex for the insect steroid hormone, 20-hydroxyecdysone (20E), is a heterodimer of EcR and USP. It has been shown that Drosophila EcR and USP can be sumoylated in mammalian cells, but it is unknown whether EcR-USP sumoylation naturally occurs in Drosophila. In Drosophila cells, USP, but not EcR, was sumoylated by Smt3, the only Drosophila SUMO protein. The presence of EcR enhanced USP sumoylation, which is further enhanced by 20E treatment. In addition to the Lys20 sumoylation site, five potential acceptor lysine residues in USP were predicted and verified. Mutation of the USP sumoylation sites or reduction of smt3 expression by RNAi attenuated 20E-induced reporter activity. Moreover, in the salivary glands, reducing smt3 expression by RNAi decreased 20E-induced reporter activity, gene expression, and autolysosome formation. Importantly, at least partially, the smt3 RNAi-mediated reduction in 20E signaling resulted from decreased protein levels of USP. In conclusion, sumoylation modulates 20E signaling by maintaining USP protein levels in Drosophila.
      Graphical abstract image

      PubDate: 2015-01-19T18:28:59Z
       
  • Mode of action of allatostatins in the regulation of juvenile hormone
           biosynthesis in the cockroach, Diploptera punctata
    • Abstract: Publication date: November 2014
      Source:Insect Biochemistry and Molecular Biology, Volume 54
      Author(s): Juan Huang , Elisabeth Marchal , Ekaterina F. Hult , Sven Zels , Jozef Vanden Broeck , Stephen S. Tobe
      The FGLamide allatostatins (FGL/ASTs) are a family of neuropeptides with pleiotropic functions, including the inhibition of juvenile hormone (JH) biosynthesis, vitellogenesis and muscle contraction. In the cockroach, Diploptera punctata, thirteen FGLa/ASTs and one allatostatin receptor (AstR) have been identified. However, the mode of action of ASTs in regulation of JH biosynthesis remains unclear. Here, we determined the tissue distribution of Dippu-AstR. And we expressed Dippu-AstR in vertebrate cell lines, and activated the receptor with the Dippu-ASTs. Our results show that all thirteen ASTs activated Dippu-AstR in a dose dependent manner, albeit with different potencies. Functional analysis of AstR in multiple cell lines demonstrated that activation of the AstR receptor resulted in elevated levels of Ca2+ and cAMP, which suggests that Dippu-AstR can act through the Gαq and Gαs protein pathways. The study on the target of AST action reveals that FGL/AST affects JH biosynthesis prior to the entry of acetyl-CoA into the JH biosynthetic pathway.
      Graphical abstract image

      PubDate: 2015-01-19T18:28:59Z
       
  • Peroxinectin catalyzed dityrosine crosslinking in the adhesive underwater
           silk of a casemaker caddisfly larvae, Hysperophylax occidentalis
    • Abstract: Publication date: November 2014
      Source:Insect Biochemistry and Molecular Biology, Volume 54
      Author(s): Ching-Shuen Wang , Nicholas N. Ashton , Robert B. Weiss , Russell J. Stewart
      Aquatic caddisfly larvae use sticky silk fibers as an adhesive tape to construct protective composite structures under water. Three new silk fiber components were identified by transcriptome and proteome analysis of the silk gland: a heme-peroxidase in the peroxinectin (Pxt) sub-family, a superoxide dismutase 3 (SOD3) that generates the H2O2 substrate of the silk fiber Pxt from environmental reactive oxygen species (eROS), and a novel structural component with sequence similarity to the elastic PEVK region of the muscle protein, titin. All three proteins are co-drawn with fibroins to form silk fibers. The Pxt and SOD3 enzymes retain activity in drawn fibers. In native fibers, Pxt activity and dityrosine crosslinks are co-localized at the boundary of a peripheral layer and the silk fiber core. To our knowledge, dityrosine crosslinks, heme peroxidase, and SOD3 activities have not been previously reported in an insect silk. The PEVK-like protein is homogeneously distributed throughout the fiber core. The results are consolidated into a model in which caddisfly silk Pxt-catalyzed dityrosine crosslinking occurs post-draw using H2O2 generated within the silk fibers by SOD3. The ROS substrate of caddisfly silk SOD3 occurs naturally in aquatic environments, from biotic and abiotic sources. The radially inhomogeneous dityrosine crosslinking and a potential titin-like PEVK protein network have important implications for the mechanical properties of caddifly silk fibers.
      Graphical abstract image

      PubDate: 2015-01-19T18:28:59Z
       
  • Allelic-specific expression in relation to Bombyx mori resistance to Bt
           toxin
    • Abstract: Publication date: November 2014
      Source:Insect Biochemistry and Molecular Biology, Volume 54
      Author(s): Yazhou Chen , Muwang Li , Iftakher Islam , Lang You , Yueqiang Wang , Zhiqian Li , Lin Ling , Baosheng Zeng , Jun Xu , Yongping Huang , Anjiang Tan
      Understanding the mechanism of Bt resistance is one of the key elements of the effective application of Bt in pest control. The lepidopteran model insect, the silkworm, demonstrates qualities that make it an ideal species to use in achieving this understanding. We screened 45 strains of silkworm (Bombyx mori) using a Cry1Ab toxin variant. The sensitivity levels of the strains varied over a wide range. A resistant strain (P50) and a phylogenetically related susceptible strain (Dazao) were selected to profile the expressions of 12 Bt resistance-related genes. The SNPs in these genes were detected based on EST analysis and were validated by allelic-specific PCR. A comparison of allelic-specific expression between P50 and Dazao showed that the transcript levels of heterozygous genes containing two alleles rather than an imbalanced allelic expression contribute more to the resistance of P50 against Bt. The responses of the allelic-specific expression to Bt in hybrid larvae were then investigated. The results showed that the gene expression pattern of an ATP-binding cassette transporter C2 (ABCC2) and an aminopeptidase N (APN3), changed in an allelic-specific manner, with the increase of the resistant allele expression correlated with larval survival. The results suggest that a trans-regulatory mechanism in ABCC2 and APN3 allelic-specific expression is involved in the insect's response to the Bt toxin. The potential role of allelic-specific gene regulation in insect resistance to Bt toxins is discussed.
      Graphical abstract image

      PubDate: 2015-01-19T18:28:59Z
       
  • CYP18A1 regulates tissue-specific steroid hormone inactivation in Bombyx
           mori
    • Abstract: Publication date: November 2014
      Source:Insect Biochemistry and Molecular Biology, Volume 54
      Author(s): Zhiqian Li , Xie Ge , Lin Ling , Baosheng Zeng , Jun Xu , Abu F.M. Aslam , Lang You , Subba Reddy Palli , Yongping Huang , Anjiang Tan
      Insect development and metamorphosis are regulated by two major hormones, juvenile hormone and ecdysteroids. Despite being the key regulator of insect developmental transitions, the metabolic pathway of the primary steroid hormone, 20-hydroxyecdysone (20E), especially its inactivation pathway, is still not completely elucidated. A cytochrome P450 enzyme, CYP18A1, has been shown to play key roles in insect steroid hormone inactivation through 26-hydroxylation. Here, we identified two CYP18 (BmCYP18A1 and BmCYP18B1) orthologs in the lepidopteran model insect, Bombyx mori. Interestingly, BmCYP18A1 gene is predominantly expressed in the middle silk gland (MSG) while BmCYP18B1 expresses ubiquitously in B. mori. BmCYP18A1 is induced by 20E in vitro, suggesting its role in 20E metabolism. Using the binary Gal4/UAS transgenic system, we ectopically overexpressed BmCYP18A1 in a MSG-specific manner with a Sericin1-Gal4 (Ser-Gal4) driver or in a ubiquitous manner with an Actin3-Gal4 (A3-Gal4) driver. Ectopic overexpression of BmCYP18A1 in MSG or in all tissues resulted in developmental arrestment of transgenic animals during the final instar larval stage. The 20E titers in the transgenic animals expressing BmCYP18A1 were lower compared to the levels in the control animals. Although the biological significance of MSG-specific expression of BmCYP18A1 is unclear, our results provide the first evidence that BmCYP18A1, which is conserved in most arthropods, is involved in a tissue-specific steroid hormone inactivation in B. mori.
      Graphical abstract image

      PubDate: 2015-01-19T18:28:59Z
       
  • Expression and characterization of an epoxide hydrolase from Anopheles
           gambiae with high activity on epoxy fatty acids
    • Abstract: Publication date: November 2014
      Source:Insect Biochemistry and Molecular Biology, Volume 54
      Author(s): Jiawen Xu , Christophe Morisseau , Bruce D. Hammock
      In insects, epoxide hydrolases (EHs) play critical roles in the metabolism of xenobiotic epoxides from the food resources and in the regulation of endogenous chemical mediators, such as juvenile hormones. Using the baculovirus expression system, we expressed and characterized an epoxide hydrolase from Anopheles gambiae (AgEH) that is distinct in evolutionary history from insect juvenile hormone epoxide hydrolases (JHEHs). We partially purified the enzyme by ion exchange chromatography and isoelectric focusing. The experimentally determined molecular weight and pI were estimated to be 35 kD and 6.3 respectively, different than the theoretical ones. The AgEH had the greatest activity on long chain epoxy fatty acids such as 14,15-epoxyeicosatrienoic acids (14,15-EET) and 9,10-epoxy-12Z-octadecenoic acids (9,10-EpOME or leukotoxin) among the substrates evaluated. Juvenile hormone III, a terpenoid insect growth regulator, was the next best substrate tested. The AgEH showed kinetics comparable to the mammalian soluble epoxide hydrolases, and the activity could be inhibited by AUDA [12-(3-adamantan-1-yl-ureido) dodecanoic acid], a urea-based inhibitor designed to inhibit the mammalian soluble epoxide hydrolases. The rabbit serum generated against the soluble epoxide hydrolase of Mus musculus can both cross-react with natural and denatured forms of the AgEH, suggesting immunologically they are similar. The study suggests there are mammalian sEH homologs in insects, and epoxy fatty acids may be important chemical mediators in insects.
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      PubDate: 2015-01-19T18:28:59Z
       
  • A novel method to study insect olfactory receptor function using
           HEK293 cells
    • Abstract: Publication date: November 2014
      Source:Insect Biochemistry and Molecular Biology, Volume 54
      Author(s): Jacob A. Corcoran , Melissa D. Jordan , Colm Carraher , Richard D. Newcomb
      The development of rapid and reliable assays to characterize insect odorant receptors (ORs) and pheromone receptors (PRs) remains a challenge for the field. Typically ORs and PRs are functionally characterized either in vivo in transgenic Drosophila or in vitro through expression in Xenopus oocytes. While these approaches have succeeded, they are not well suited for high-throughput screening campaigns, primarily due to inherent characteristics that limit their ability to screen large quantities of compounds in a short period of time. The development of a practical, robust and consistent in vitro assay for functional studies on ORs and PRs would allow for high-throughput screening for ligands, as well as for compounds that could be used as novel olfactory-based pest management tools. Here we describe a novel method of utilizing human embryonic kidney cells (HEK293) transfected with inducible receptor constructs for the functional characterization of ORs in 96-well plates using a fluorescent spectrophotometer. Using EposOrco and EposOR3 from the pest moth, Epiphyas postvittana as an example, we generated HEK293 cell lines with robust and consistent responses to ligands in functional assays. Single-cell sorting of cell lines by FACS facilitated the selection of isogenic cell lines with maximal responses, and the addition of epitope tags on the N-termini allowed the detection of recombinant proteins in homogenates by western blot and in cells by immunocytochemistry. We thoroughly describe the methods used to generate these OR-expressing cell lines, demonstrating that they have all the necessary features required for use in high-throughput screening platforms.
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      PubDate: 2015-01-19T18:28:59Z
       
  • Regulation of the gut-specific carboxypeptidase: A study using the binary
           Gal4/UAS system in the mosquito Aedes aegypti
    • Abstract: Publication date: November 2014
      Source:Insect Biochemistry and Molecular Biology, Volume 54
      Author(s): Bo Zhao , Vladimir A. Kokoza , Tusar T. Saha , Stephanie Wang , Sourav Roy , Alexander S. Raikhel
      Pathogen transmission by mosquitoes is tightly linked to blood feeding which, in turn, is required for egg development. Studies of these processes would greatly benefit from genetic methods, such as the binary Gal4/UAS system. The latter has been well established for model organisms, but its availability is limited for mosquitoes. The objective of this study was to develop the blood-meal-activated, gut-specific Gal4/UAS system for the yellow-fever mosquito Aedes aegypti and utilize it to investigate the regulation of gut-specific gene expression. A 1.1-kb, 5′ upstream region of the carboxypeptidase A (CP) gene was used to genetically engineer the CP-Gal4 driver mosquito line. The CP-Gal4 specifically activated the Enhanced Green Fluorescent Protein (EGFP) reporter only after blood feeding in the gut of the CP-Gal4 > UAS-EGFP female Ae. aegypti. We used this system to study the regulation of CP gene expression. In vitro treatments with either amino acids (AAs) or insulin stimulated expression of the CP-Gal4 > UAS-EGFP transgene; no effect was observed with 20-hydroxyecdysone (20E) treatments. The transgene activation by AAs and insulin was blocked by rapamycin, the inhibitor of the Target-of-Rapamycin (TOR) kinase. RNA interference (RNAi) silence of the insulin receptor (IR) reduced the expression of the CP-Gal4 > UAS-EGFP transgene. Thus, in vitro and in vivo experiments have revealed that insulin and TOR pathways control expression of the digestive enzyme CP. In contrast, 20E, the major regulator of post-blood-meal vitellogenic events in female mosquitoes, has no role in regulating the expression of this gene. This novel CP-Gal4/UAS system permits functional testing of midgut-specific genes that are involved in blood digestion and interaction with pathogens in Ae. aegypti mosquitoes.
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      PubDate: 2015-01-19T18:28:59Z
       
  • The role of Rdl in resistance to phenylpyrazoles in Drosophila
           melanogaster
    • Abstract: Publication date: November 2014
      Source:Insect Biochemistry and Molecular Biology, Volume 54
      Author(s): Emily J. Remnant , Craig J. Morton , Phillip J. Daborn , Christopher Lumb , Ying Ting Yang , Hooi Ling Ng , Michael W. Parker , Philip Batterham
      Extensive use of older generation insecticides may result in pre-existing cross-resistance to new chemical classes acting at the same target site. Phenylpyrazole insecticides block inhibitory neurotransmission in insects via their action on ligand-gated chloride channels (LGCCs). Phenylpyrazoles are broad-spectrum insecticides widely used in agriculture and domestic pest control. So far, all identified cases of target site resistance to phenylpyrazoles are based on mutations in the Rdl (Resistance to dieldrin) LGCC subunit, the major target site for cyclodiene insecticides. We examined the role that mutations in Rdl have on phenylpyrazole resistance in Drosophila melanogaster, exploring naturally occurring variation, and generating predicted resistance mutations by mutagenesis. Natural variation at the Rdl locus in inbred strains of D. melanogaster included gene duplication, and a line containing two Rdl mutations found in a highly resistant line of Drosophila simulans. These mutations had a moderate impact on survival following exposure to two phenylpyrazoles, fipronil and pyriprole. Homology modelling suggested that the Rdl chloride channel pore contains key residues for binding fipronil and pyriprole. Mutagenesis of these sites and assessment of resistance in vivo in transgenic lines showed that amino acid identity at the Ala301 site influenced resistance levels, with glycine showing greater survival than serine replacement. We confirm that point mutations at the Rdl 301 site provide moderate resistance to phenylpyrazoles in D. melanogaster. We also emphasize the beneficial aspects of testing predicted mutations in a whole organism to validate a candidate gene approach.
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      PubDate: 2015-01-19T18:28:59Z
       
  • Editorial Board
    • Abstract: Publication date: November 2014
      Source:Insect Biochemistry and Molecular Biology, Volume 54




      PubDate: 2015-01-19T18:28:59Z
       
  • Tweedle cuticular protein BmCPT1 is involved in innate immunity by
           participating in recognition of Escherichia coli
    • Abstract: Publication date: Available online 20 November 2014
      Source:Insect Biochemistry and Molecular Biology
      Author(s): Jiubo Liang , Ting Wang , Zhonghuai Xiang , Ningjia He
      Bombyx mori, a lepidopteran insect, is one of the earliest models for pattern recognition of Gram-negative bacteria, which may induce the IMD pathway for production of antibacterial peptides. So far, several recognition proteins have been reported in B. mori. However, the connection between pattern recognition of Gram negative bacteria and activation of BmRelish1, a transcription factor controlled by the IMD pathway remains largely unknown. In the present study, we identify BmCPT1, a cuticle protein bearing a Tweedle domain. Its gene expression is co-regulated by NF-kappaB and juvenile hormone signals. BmCPT1 is induced by Escherichia coli in fat bodies and hemocytes, but is constitutively expressed in the epidermis. In vitro binding assays indicate that BmCPT1 protein recognizes and binds to E. coli peptidoglycan. Post-transcriptionally modified BmCPT1 in the hemolymph binds to E. coli cells through interactions with peptidoglycan recognition protein-5 (BmPGRP5) and lipopolysaccharide binding protein (BmLBP). Transgenic overexpression of BmCPT1 causes the upregulated expression of BmRelish1 and clear induction of two gloverin genes. Therefore, BmCPT1 may work along with BmPGRP-S5 and BmLBP to recognize E. coli in the hemolymph and indirectly activate BmRelish1 to induce antimicrobial peptide synthesis.
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      PubDate: 2015-01-19T18:28:59Z
       
  • Involvement of FTZ-F1 in the regulation of pupation in Leptinotarsa
           decemlineata (Say)
    • Abstract: Publication date: December 2014
      Source:Insect Biochemistry and Molecular Biology, Volume 55
      Author(s): Xin-Ping Liu , Kai-Yun Fu , Feng-Gong Lü , Qing-Wei Meng , Wen-Chao Guo , Guo-Qing Li
      During the final instar larvae of holometabolous insects, a pulse of 20-hydroxyecdysone (20E) and a drop in juvenile hormone (JH) trigger larval-pupal metamorphosis. In this study, two LdFTZ-F1 cDNAs (LdFTZ-F1-1 and LdFTZ-F1-2) were cloned in Leptinotarsa decemlineata. Both LdFTZ-F1-1 and LdFTZ-F1-2 were highly expressed just before or right after each molt, similar to the expression pattern of an ecdysteroidogenesis gene LdSHD. Ingestion of an ecdysteroid agonist halofenozide (Hal) enhanced LdFTZ-F1-1 and LdFTZ-F1-2 expression in the final larval instar. Conversely, a decrease in 20E by feeding a double-stranded RNA (dsRNA) against LdSHD repressed the expression. Moreover, Hal rescued the expression levels in LdSHD-silenced larvae. Thus, 20E peaks seem to induce the transcription of LdFTZ-F1s. Furthermore, ingesting dsLdFTZ-F1 from a common fragment of LdFTZ-F1-1 and LdFTZ-F1-2 successfully knocked down both LdFTZ-F1s, and impaired pupation. Finally, knocking down LdFTZ-F1s significantly repressed the transcription of three ecdysteroidogenesis genes, lowered 20E titer, and reduced the expression of two 20E receptor genes. Silencing LdFTZ-F1s also induced the expression of a JH biosynthesis gene, increased JH titer, but decreased the mRNA level of a JH early-inducible gene. Thus, LdFTZ-F1s are involved in the regulation of pupation by modulating 20E and JH titers and mediating their signaling pathways.
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      PubDate: 2015-01-19T18:28:59Z
       
  • Re-examination of a α-chymotrypsin-solubilized laccase in the pupal
           cuticle of the silkworm, Bombyx mori: Insights into the regulation system
           for laccase activation during the ecdysis process
    • Abstract: Publication date: December 2014
      Source:Insect Biochemistry and Molecular Biology, Volume 55
      Author(s): Tsunaki Asano , Masato Taoka , Yoshio Yamauchi , R. Craig Everroad , Yosuke Seto , Toshiaki Isobe , Masaharu Kamo , Naoyuki Chosa
      The laccase in the pupal cuticle of the silkworm, Bombyx mori, is thought to accumulate as an inactive precursor that can be activated stage-dependently. In this study we isolated an 81-kDa laccase from cuticular extract of B. mori that was prepared by digestion of the pupal cuticles with α-chymotrypsin. The mass spectrometric analysis of the purified protein indicates that this 81-kDa laccase is a product of the Bombyx laccase2 gene. The purified 81-kDa laccase (α-chymotrypsin-solubilized Bombyx laccase2: Bm-clac2) has an N-terminal sequence of RNPADS that corresponds to Arg146 to Ser151 of the deduced protein sequence of Bmlaccase2 cDNA, indicating that Bm-clac2 lacks the N-terminal part upstream from residue Arg146. Bm-clac2 shows enzymatic activity, but its specific activity is increased around 17-fold after treatment with trypsin, which involves cleavage of peptide bonds at the C-terminal region. We also found that the activity of Bm-clac2 is increased in the presence of isopropanol. In previous reports, proteolytic processing has been hypothesized as a system for laccase activation in vivo, but the present result implies that this type of processing is not the only way to convert Bm-clac2 to the high-activity enzyme.
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      PubDate: 2015-01-19T18:28:59Z
       
  • Identification of differentially expressed microRNAs in Culex pipiens and
           their potential roles in pyrethroid resistance
    • Abstract: Publication date: December 2014
      Source:Insect Biochemistry and Molecular Biology, Volume 55
      Author(s): Shanchao Hong , Qin Guo , Weijie Wang , Shengli Hu , Fujin Fang , Yuan Lv , Jing Yu , Feifei Zou , Zhentao Lei , Kai Ma , Lei Ma , Dan Zhou , Yan Sun , Donghui Zhang , Bo Shen , Changliang Zhu
      Pyrethroids are the major class of insecticides used for mosquito control. Excessive and improper use of insecticides, however, has resulted in pyrethroid resistance, which has become a major obstacle for mosquito control. The development of pyrethroid resistance is a complex process involving many genes, and information on post-transcription regulation of pyrethroid resistance is lacking. In this study, we extracted RNA from mosquitoes in various life stages (fourth-instar larvae, pupae, male and female adult mosquitoes) from deltamethrin-sensitive (DS) and resistant (DR) strains. Using illumina sequencing, we obtained 13760296 and 12355472 reads for DS-strains and DR-strains, respectively. We identified 100 conserved miRNAs and 42 novel miRNAs derived from 21 miRNA precursors in Culex pipiens. After normalization, we identified 28 differentially expressed miRNAs between the two strains. Additionally, we found that cpp-miR-71 was significant down regulated in female adults from the DR-strain. Based on microinjection and CDC Bottle Bioassay data, we found that cpp-miR-71 may play a contributing role in deltamethrin resistance. The present study provides the firstly large-scale characterization of miRNAs in Cu. pipiens and provides evidence of post-transcription regulation. The differentially expressed miRNAs between the two strains are expected to contribute to the development of pyrethroid resistance.
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      PubDate: 2015-01-19T18:28:59Z
       
  • Site-specific, TALENs-mediated transformation of Bombyx mori
    • Abstract: Publication date: December 2014
      Source:Insect Biochemistry and Molecular Biology, Volume 55
      Author(s): Yueqiang Wang , Anjiang Tan , Jun Xu , Zhiqian Li , Baosheng Zeng , Lin Ling , Lang You , Yazhou Chen , Anthony A. James , Yongping Huang
      Transposon-based genetic transformation has facilitated insect functional genomics and new strategies of pest management. However, there is a need for alternative, site-specific approaches to overcome limitations of random integration (and associated position-effects) and potential instability of inserted transgenes. Here we describe a transposon-free, site-specific genetic transformation system mediated by transcription activator-like effector nucleases (TALENs) in the silkworm, Bombyx mori, a lepidopteran model insect. We successfully established a site-specific transgenic system with comparable transformation efficiency to transposon-based genetic transformation through microinjection of TALENs mRNA targeting the BmBLOS2 locus and a linearizable donor plasmid encoding an expression cassette of the DsRed2 red fluorescent protein. This system provides a valuable approach for insect transgenesis and will enable future functional gene analysis and generate novel applications in agricultural and medical insect pest-management technologies.
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      PubDate: 2015-01-19T18:28:59Z
       
  • A novel β-fructofuranosidase in Coleoptera: Characterization of a
           β-fructofuranosidase from the sugarcane weevil, Sphenophorus levis
    • Abstract: Publication date: December 2014
      Source:Insect Biochemistry and Molecular Biology, Volume 55
      Author(s): Rafael Pedezzi , Fernando P.P. Fonseca , Célio Dias Santos Júnior , Luciano T. Kishi , Walter R. Terra , Flávio Henrique-Silva
      β-fructofuranosidases or invertases (EC 3.2.1.26) catalyze the hydrolysis of sucrose into fructose and glucose. β-fructofuranosidases have been widely described in microorganisms, but were not known in the animal kingdom until very recently. There are studies reporting lepidopteran β-fructofuranosidases, but no β-fructofuranosidase gene sequence or encoding transcript has previously been identified in beetles. Considering the scarcity of functional studies on insect β-fructofuranosidases and their apparent non-occurrence among coleopterans, the aim of the present study was to investigate the occurrence and characterize a β-fructofuranosidase transcript identified in a cDNA library from the sugarcane weevil, Sphenophorus levis (Curculionidae). To validate that the β-fructofuranosidase sequence (herein denominated Sl-β-fruct) is indeed encoded by the S. levis genome, PCRs were performed using genomic DNA extracted from the larval fat body as well as DNA from the midgut with microbial content. Amplification of Sl-β-fruct gene using larval fat body DNA indicated its presence in the insect's genomic DNA. The Sl-β-fruct gene was cloned in Pichia pastoris to produce the recombinant enzyme (rSl-β-fruct). Molecular weight of the recombinant protein was about 64 kDa, indicating possible glycosylation, since the theoretical weight was 54.8 kDa. The substrate specificity test revealed that rSl-β-fruct hydrolyzes sucrose and raffinose, but not melibiose or maltose, thereby confirming invertase activity. The pH curve revealed greatest activity at pH 5.0, demonstrating rSl-β-fruct to be an acidic β-fructofuranosidase. Quantitative PCR (qRT-PCR) analyses indicated that the production of mRNA only occurs in the midgut and reaches the greatest expression level in 30-day-old larvae, which is the expected pattern for digestive enzymes. Chromatography of glycosidases from S. levis midguts showed two enzymes acting as β-fructofuranosidase, indicating the presence of a Sl-β-fruct isoform or a β-fructofuranosidase from insect intestinal microbiota. Moreover, it was found that α-glucosidases do not act on sucrose hydrolysis. Phylogenetic analyses indicated this enzyme to be similar to enzymes found in other coleopteran and lepidopteran β-fructofuranosidases, but also closely similar to bacterial enzymes, suggesting potential horizontal gene transfer. Despite this, the enzyme seems to be restricted to different groups of bacteria, which suggests distinct origin events. The present study expands the concept of the occurrence of β-fructofuranosidase in insects. Despite the few descriptions of this gene in the animal kingdom, it is possible to state that β-fructofuranosidase is crucial to the establishment of some insects throughout their evolutionary history, especially members of the Lepidoptera and Coleoptera clades.
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      PubDate: 2015-01-19T18:28:59Z
       
  • Acp70A regulates Drosophila pheromones through juvenile hormone induction
    • Abstract: Publication date: January 2015
      Source:Insect Biochemistry and Molecular Biology, Volume 56
      Author(s): Gwénaëlle Bontonou , Haq Abdul Shaik , Béatrice Denis , Claude Wicker-Thomas
      Mated Drosophila melanogaster females show a decrease in mating receptivity, enhanced ovogenesis, egg-laying and activation of juvenile hormone (JH) production. Components in the male seminal fluid, especially the sex peptide ACP70A stimulate these responses in females. Here we demonstrate that ACP70A is involved in the down-regulation of female sex pheromones and hydrocarbon (CHC) production. Drosophila G10 females which express Acp70A under the control of the vitellogenin gene yp1, produced fewer pheromones and CHCs. There was a dose-dependent relationship between the number of yp1-Acp70A alleles and the reduction of these compounds. Similarly, a decrease in CHCs and diene pheromones was observed in da > Acp70A flies that ubiquitously overexpress Acp70A. Quantitative-PCR experiments showed that the expression of Acp70A in G10 females was the same as in control males and 5 times lower than in da > Acp70A females. Three to four days after injection with 4.8 pmol ACP70A, females from two different strains, exhibited a significant decrease in CHC and pheromone levels. Similar phenotypes were observed in ACP70A injected flies whose ACP70A receptor expression was knocked-down by RNAi and in flies which overexpress ACP70A N-terminal domain. These results suggest that the action of ACP70A on CHCs could be a consequence of JH activation. Female flies exposed to a JH analog had reduced amounts of pheromones, whereas genetic ablation of the corpora allata or knock-down of the JH receptor Met, resulted in higher amounts of both CHCs and pheromonal dienes. Mating had negligible effects on CHC levels, however pheromone amounts were slightly reduced 3 and 4 days post copulation. The physiological significance of ACP70A on female pheromone synthesis is discussed.
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      PubDate: 2015-01-19T18:28:59Z
       
  • Dynamics of polycomb proteins-mediated histone modifications during UV
           irradiation-induced DNA damage
    • Abstract: Publication date: December 2014
      Source:Insect Biochemistry and Molecular Biology, Volume 55
      Author(s): Zhiqing Li , Hiroaki Mon , Hitoshi Mitsunobu , Li Zhu , Jian Xu , Jae Man Lee , Takahiro Kusakabe
      Polycomb group (PcG) complexes are known to be chromatin modifiers and transcriptional repressors. In this work, we reported that the histone-modifying PcG complexes are able to participate in the repair process of ultraviolet (UV)-induced DNA lesions in the silkworm, Bombyx mori. The silkworm cells with depletion of PcG genes showed hypersensitive to UV–C irradiation and increased inhibition of cell proliferation. Interestingly, an SQ site in the silkworm-human chimeric H2A protein synthesized here was phosphorylated rapidly upon UV–C exposure, which could be used as a marker for monitoring the response to DNA damage in silkworm cells. Under these UV–C irradiated conditions, we found that PRC1-mediated ubiquitylation of H2AX, but not of H2AZ, were decreased and this deubiquitylation was independent of its phosphorylation event. In contrast, UV–C irradiation induced the increase of trimethylation of lysine 27 on histone H3 (H3K27me3), a mark of transcriptionally silent chromatin catalyzed by another PcG subcomplex, PRC2. Collectively, we provided the first evidence on chromatin remodeling in response to UV–C lesion in silkworm and revealed another layer role for PcG complexes-mediated histone modifications in contributing to creating an open chromatin structure for the efficient repair of DNA damages.
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      PubDate: 2015-01-19T18:28:59Z
       
  • The vacuolar protein sorting genes in insects: A comparative
           genome view
    • Abstract: Publication date: Available online 5 December 2014
      Source:Insect Biochemistry and Molecular Biology
      Author(s): Zhaofei Li , Gary Blissard
      In eukaryotic cells, regulated vesicular trafficking is critical for directing protein transport and for recycling and degradation of membrane lipids and proteins. Through carefully regulated transport vesicles, the endomembrane system performs a large and important array of dynamic cellular functions while maintaining the integrity of the cellular membrane system. Genetic studies in yeast Saccharomyces cerevisiae have identified approximately 50 vacuolar protein sorting (VPS) genes involved in vesicle trafficking, and most of these genes are also characterized in mammals. The VPS proteins form distinct functional complexes, which include complexes known as ESCRT, retromer, CORVET, HOPS, GARP, and PI3K-III. Little is known about the orthologs of VPS proteins in insects. Here, with the newly annotated Manduca sexta genome, we carried out genomic comparative analysis of VPS proteins in yeast, humans, and 13 sequenced insect genomes representing the Orders Hymenoptera, Diptera, Hemiptera, Phthiraptera, Lepidoptera, and Coleoptera. Amino acid sequence alignments and domain/motif structure analyses reveal that most of the components of ESCRT, retromer, CORVET, HOPS, GARP, and PI3K-III are evolutionarily conserved across yeast, insects, and humans. However, in contrast to the VPS gene expansions observed in the human genome, only four VPS genes (VPS13, VPS16, VPS33, and VPS37) were expanded in the six insect Orders. Additionally, VPS2 was expanded only in species from Phthiraptera, Lepidoptera, and Coleoptera. These studies provide a baseline for understanding the evolution of vesicular trafficking across yeast, insect, and human genomes, and also provide a basis for further addressing specific functional roles of VPS proteins in insects.
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      PubDate: 2015-01-19T18:28:59Z
       
  • Editorial Board
    • Abstract: Publication date: December 2014
      Source:Insect Biochemistry and Molecular Biology, Volume 55




      PubDate: 2015-01-19T18:28:59Z
       
  • The cyclic keto-enol insecticide spirotetramat inhibits insect and spider
           mite acetyl-CoA carboxylases by interfering with the carboxyltransferase
           partial reaction
    • Abstract: Publication date: December 2014
      Source:Insect Biochemistry and Molecular Biology, Volume 55
      Author(s): Peter Lümmen , Jahangir Khajehali , Kai Luther , Thomas Van Leeuwen
      Acetyl-CoA carboxylase (ACC) catalyzes the committed and rate-limiting step in fatty acid biosynthesis. The two partial reactions, carboxylation of biotin followed by carboxyl transfer to the acceptor acetyl-CoA, are performed by two separate domains in animal ACCs. The cyclic keto-enol insecticides and acaricides have been proposed to inhibit insect ACCs. In this communication, we show that the enol derivative of the cylic keto-enol insecticide spirotetramat inhibited ACCs partially purified from the insect species Myzus persicae and Spodoptera frugiperda, as well as the spider mite (Tetranychus urticae) ACC which was expressed in insect cells using a recombinant baculovirus. Steady-state kinetic analysis revealed competitive inhibition with respect to the carboxyl acceptor, acetyl-CoA, indicating that spirotetramat-enol bound to the carboxyltransferase domain of ACC. Interestingly, inhibition with respect to the biotin carboxylase substrate ATP was uncompetitive. Amino acid residues in the carboxyltransferase domains of plant ACCs are important for binding of established herbicidal inhibitors. Mutating the spider mite ACC at the homologous positions, for example L1736 to either isoleucine or alanine, and A1739 to either valine or serine, did not affect the inhibition of the spider mite ACC by spirotetramat-enol. These results indicated different binding modes of the keto-enols and the herbicidal chemical families.
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      PubDate: 2015-01-19T18:28:59Z
       
  • Expression and evolution of hexamerins from the tobacco hornworm, Manduca
           sexta, and other Lepidoptera
    • Abstract: Publication date: Available online 8 December 2014
      Source:Insect Biochemistry and Molecular Biology
      Author(s): Thorsten Burmester
      Hexamerins are large hemolymph-proteins that accumulate during the late larval stages of insects. Hexamerins have emerged from hemocyanin, but have lost the ability to bind oxygen. Hexamerins are mainly considered as storage proteins for non-feeding stages, but may also have other functions, e.g. in cuticle formation, transport and immune response. The genome of the hornworm Manduca sexta harbors six hexamerin genes. Two of them code for arylphorins (Msex2.01690, Msex2.15504) and two genes correspond to a methionine-rich hexamerin (Msex2.10735) and a moderately methionine-rich hexamerin (Msex2.01694), respectively. Two other genes do not correspond to any known hexamerin and distantly resemble the arylphorins (Msex2.01691, Msex2.01693). Five of the six hexamerin genes are clustered within ∼45 kb on scaffold 00023, which shows conserved synteny in various lepidopteran genomes. The methionine-rich hexamerin gene is located at a distinct site. M. sexta and other Lepidoptera have lost the riboflavin-binding hexamerin. With the exception of Msex2.01691, which displays low mRNA levels throughout the life cycle, all hexamerins are most highly expressed during pre-wandering phase of the 5th larval instar of M. sexta, supporting their role as storage proteins. Notably, Msex2.01691 is most highly expressed in the brain, suggesting a divergent function. Phylogenetic analyses showed that hexamerin evolution basically follows insect systematics. Lepidoptera display an unparalleled diversity of hexamerins, which exceeds that of other hexapod orders. In contrast to previous analyses, the lepidopteran hexamerins were found monophyletic. Five distinct types of hexamerins have been identified in this order, which differ in terms of amino acid composition and evolutionary history: i. the arylphorins, which are rich in aromatic amino acids (∼20% phenylalanine and tyrosine), ii. the distantly related arylphorin-like hexamerins, iii. the methionine-rich hexamerins, iv. the moderately methionine rich hexamerins, and v. the riboflavin-binding hexamerins.
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      PubDate: 2015-01-19T18:28:59Z
       
  • Analysis of chitin-binding proteins from Manduca sexta provides new
           insights into evolution of peritrophin A-type chitin-binding domains in
           insects
    • Abstract: Publication date: Available online 15 December 2014
      Source:Insect Biochemistry and Molecular Biology
      Author(s): Guillaume Tetreau , Neal T. Dittmer , Xiaolong Cao , Sinu Agrawal , Yun-Ru Chen , Subbaratnam Muthukrishnan , Jiang Haobo , Gary W. Blissard , Michael R. Kanost , Ping Wang
      In insects, chitin is a major structural component of the cuticle and the peritrophic membrane (PM). In nature, chitin is always associated with proteins among which chitin-binding proteins (CBPs) are the most important for forming, maintaining and regulating the functions of these extracellular structures. In this study, a genome-wide search for genes encoding proteins with ChtBD2-type (peritrophin A-type) chitin-binding domains (CBDs) was conducted. A total of 53 genes encoding 56 CBPs were identified, including 15 CPAP1s (cuticular proteins analogous to peritrophins with 1 CBD), 11 CPAP3s (CPAPs with 3 CBDs) and 17 PMPs (PM proteins) with a variable number of CBDs, which are structural components of cuticle or of the PM. CBDs were also identified in enzymes of chitin metabolism including 6 chitinases and 7 chitin deacetylases encoded by 6 and 5 genes, respectively. RNA-seq analysis confirmed that PMP and CPAP genes have differential spatial expression patterns. The expression of PMP genes is midgut-specific, while CPAP genes are widely expressed in different cuticle forming tissues. Phylogenetic analysis of CBDs of proteins in insects belonging to different orders revealed that CPAP1s from different species constitute a separate family with 16 different groups, including 6 new groups identified in this study. The CPAP3s are clustered into a separate family of 7 groups present in all insect orders. Altogether, they reveal that duplication events of CBDs in CPAP1s and CPAP3s occurred prior to the evolutionary radiation of insect species. In contrast to the CPAPs, all CBDs from individual PMPs are generally clustered and distinct from other PMPs in the same species in phylogenetic analyses, indicating that the duplication of CBDs in each of these PMPs occurred after divergence of insect species. Phylogenetic analysis of these three CBP families showed that the CBDs in CPAP1s form a clearly separate family, while those found in PMPs and CPAP3s were clustered together in the phylogenetic tree. For chitinases and chitin deacetylases, most of phylogenetic analysis performed with the CBD sequences resulted in similar clustering to the one obtained by using catalytic domain sequences alone, suggesting that CBDs were incorporated into these enzymes and evolved in tandem with the catalytic domains before the diversification of different insect orders. Based on these results, the evolution of CBDs in insect CBPs is discussed to provide a new insight into the CBD sequence structure and diversity, and their evolution and expression in insects.
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      PubDate: 2015-01-19T18:28:59Z
       
  • Editorial Board
    • Abstract: Publication date: February 2015
      Source:Insect Biochemistry and Molecular Biology, Volume 57




      PubDate: 2015-01-19T18:28:59Z
       
  • Identification and functional characterization of FGLamide-related
           allatostatin receptor in Rhodnius prolixus
    • Abstract: Publication date: February 2015
      Source:Insect Biochemistry and Molecular Biology, Volume 57
      Author(s): Meet Zandawala , Ian Orchard
      FGLamide-related ASTs (FGLa/ASTs) are a family of brain/gut peptides with numerous physiological roles, including inhibition of juvenile hormone (JH) biosynthesis by the corpora allata and inhibition of visceral muscle contraction. FGLa/ASTs mediate their effects by binding to a rhodopsin-like G-protein coupled receptor that is evolutionarily related to the vertebrate galanin receptor. Here we determine the cDNA sequence encoding FGLa/AST receptor (FGLa/AST-R) from the Chagas disease vector, Rhodnius prolixus (Rhopr-FGLa/AST-R), determine its spatial expression pattern using quantitative PCR and functionally characterize the receptor using a heterologous assay. Our expression analysis indicates that Rhopr-FGLa/AST-R is highly expressed in the central nervous system. The receptor is also expressed in various peripheral tissues including the dorsal vessel, midgut, hindgut and reproductive tissues of both males and females, suggesting a role in processes associated with feeding and reproduction. The possible involvement of Rhopr-FGLa/ASTs in the inhibition of JH biosynthesis is also implicated due to presence of the receptor transcript in the R. prolixus corpora cardiaca/corpora allata complex. The functional assay showed that various Rhopr-FGLa/ASTs activate the receptor, with EC50 values for the response in the nanomolar range. Moreover, Rhopr-FGLa/AST-R can couple with Gq alpha subunits and cause an increase in intracellular calcium concentration. Lastly, we tested various FGLa/AST analogs in our heterologous assay. These compounds also activated the receptor and thus have the potential to serve as insect growth regulators and aid in pest control.


      PubDate: 2015-01-19T18:28:59Z
       
  • LIM-homeodomain transcription factor Awh is a key component activating all
           three fibroin genes, fibH, fibL and fhx, in the silk gland of the
           silkworm, Bombyx mori
    • Abstract: Publication date: January 2015
      Source:Insect Biochemistry and Molecular Biology, Volume 56
      Author(s): Mai Kimoto , Takuya Tsubota , Keiro Uchino , Hideki Sezutsu , Shigeharu Takiya
      In the silkworm Bombyx mori, three fibroin genes, fibroin-heavy-chain (fibH), fibroin-light-chain (fibL) and fibrohexamerin (fhx), are coexpressed only in the posterior silk gland (PSG) cells, while the sericin genes encoding silk glue proteins are expressed in the middle silk gland (MSG) cells. Silk gland factor-2 (SGF-2) is a PSG-specific activator complex of fibH, composed of a LIM-homeodomain protein, Awh, and its cofactors, Ldb and Lcaf. We investigated whether SGF-2 can activate other fibroin genes using transgenic silkworms. The genes for Ldb and Lcaf were expressed ubiquitously in various tissues, while the gene for Awh was expressed strictly specific in PSG of the wild type silkworms. Misexpression of Awh in transgenic silkworms induced ectopic expression of fibL and fhx as well as fibH in MSG. Coincidently with the induction of fibL and fhx by Awh, binding of SGF-2 to the promoter of fibL and fhx was detected in vitro, and SGF-2 binds directly to the fhx core promoter. Ectopic expression of the fibroin genes was observed at high levels in the middle part of MSG. Moreover, fibL and fhx were induced in the anterior silk gland (ASG) of the transgenic silkworms, but fibH was not. These results indicate that Awh is a key activator of all three fibroin genes, and the activity is probably regulated in conjunction with additional factors.
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      PubDate: 2015-01-19T18:28:59Z
       
  • Pharmacological and signalling properties of a D2-like dopamine receptor
           (Dop3) in Tribolium castaneum
    • Abstract: Publication date: January 2015
      Source:Insect Biochemistry and Molecular Biology, Volume 56
      Author(s): Heleen Verlinden , Rut Vleugels , Rik Verdonck , Elodie Urlacher , Jozef Vanden Broeck , Alison Mercer
      Dopamine is an important neurotransmitter in the central nervous system of vertebrates and invertebrates. Despite their evolutionary distance, striking parallels exist between deuterostomian and protostomian dopaminergic systems. In both, signalling is achieved via a complement of functionally distinct dopamine receptors. In this study, we investigated the sequence, pharmacology and tissue distribution of a D2-like dopamine receptor from the red flour beetle Tribolium castaneum (TricaDop3) and compared it with related G protein-coupled receptors in other invertebrate species. The TricaDop3 receptor-encoding cDNA shows considerable sequence similarity with members of the Dop3 receptor class. Real time qRT-PCR showed high expression in both the central brain and the optic lobes, consistent with the role of dopamine as neurotransmitter. Activation of TricaDop3 expressed in mammalian cells increased intracellular Ca2+ signalling and decreased NKH-477 (a forskolin analogue)-stimulated cyclic AMP levels in a dose-dependent manner. We studied the pharmacological profile of the TricaDop3 receptor and demonstrated that the synthetic vertebrate dopamine receptor agonists, 2 – amino- 6,7 – dihydroxy – 1,2,3,4 – tetrahydronaphthalene hydrobromide (6,7-ADTN) and bromocriptine acted as agonists. Methysergide was the most potent of the antagonists tested and showed competitive inhibition in the presence of dopamine. This study offers important information on the Dop3 receptor from Tribolium castaneum that will facilitate functional analyses of dopamine receptors in insects and other invertebrates.
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      PubDate: 2015-01-19T18:28:59Z
       
  • Chitin is a necessary component to maintain the barrier function of the
           peritrophic matrix in the insect midgut
    • Abstract: Publication date: January 2015
      Source:Insect Biochemistry and Molecular Biology, Volume 56
      Author(s): Marco Kelkenberg , Jothini Odman-Naresh , Subbaratnam Muthukrishnan , Hans Merzendorfer
      In most insects, the peritrophic matrix (PM) partitions the midgut into different digestive compartments, and functions as a protective barrier against abrasive particles and microbial infections. In a previous study we demonstrated that certain PM proteins are essential in maintaining the PM's barrier function and establishing a gradient of PM permeability from the anterior to the posterior part of the midgut which facilitates digestion (Agrawal et al., 2014). In this study, we focused on the effects of a reduction in chitin content on PM permeability in larvae of the red flour beetle, Tribolium castaneum. Oral administration of the chitin synthesis inhibitor diflubenzuron (DFB) only partially reduced chitin content of the larval PM even at high concentrations. We observed no nutritional effects, as larval growth was unaffected and neutral lipids were not depleted from the fat body. However, the metamorphic molt was disrupted and the insects died at the pharate pupal stage, presumably due to DFB's effect on cuticle formation. RNAi to knock-down expression of the gene encoding chitin synthase 2 in T. castaneum (TcCHS-2) caused a complete loss of chitin in the PM. Larval growth was significantly reduced, and the fat body was depleted of neutral lipids. In situ PM permeability assays monitoring the distribution of FITC dextrans after DFB exposure or RNAi for TcCHS-2 revealed that PM permeability was increased in both cases. RNAi for TcCHS-2, however, led to a higher permeation of the PM by FITC dextrans than DFB treatment even at high doses. Similar effects were observed when the chitin content was reduced by feeding DFB to adult yellow fever mosquitos, Aedes aegypti. We demonstrate that the presence of chitin is necessary for maintaining the PM's barrier function in insects. It seems that the insecticidal effects of DFB are mediated by the disruption of cuticle synthesis during the metamorphic molt rather than by interfering with larval nutrition. However, as DFB clearly affects PM permeability, it may be suitable to increase the efficiency of pesticides targeting the midgut.
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      PubDate: 2015-01-19T18:28:59Z
       
  • Editorial Board
    • Abstract: Publication date: January 2015
      Source:Insect Biochemistry and Molecular Biology, Volume 56




      PubDate: 2015-01-19T18:28:59Z
       
  • Syntaxin 1A modulates the sexual maturity rate and progeny egg size
           related to phase changes in locusts
    • Abstract: Publication date: January 2015
      Source:Insect Biochemistry and Molecular Biology, Volume 56
      Author(s): Qianquan Chen , Jing He , Chuan Ma , Dan Yu , Le Kang
      The migratory locust (Locusta migratoria) exhibits clear phenotypic plasticity depending on its population density. Previous studies have explored the molecular mechanisms of body colour, behavior, immunity, and metabolism between high population density gregarious (G) and low population density solitarious (S) locusts. However, the molecular mechanisms underlying differences in reproductive traits remain unknown. G locusts reach sexual maturation much faster and lay larger eggs compared with S locusts. The traits of G locusts decreased significantly with isolation, whereas those of S locusts increased with crowding. Analysis of gene expression in female adults indicated that syntaxin 1A (Syx1A) was expressed significantly higher in G locusts than in S locusts. After silencing Syx1A expression in G locusts by RNA interference (RNAi), their sexual maturity rate and progeny egg size changed towards those of S locusts. Similarly, increment in the traits of S locusts with crowding was blocked by Syx1A interference. Changes in the traits were also confirmed by decrease in the level of vitellogenin, which is regulated by Syx1A. In conclusion, plasticity of the sexual maturity rate and progeny egg size of G and S locusts, which is beneficial for locusts to adapt to environmental changes, is regulated by Syx1A.
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      PubDate: 2015-01-19T18:28:59Z
       
  • Annotation and expression analysis of cuticular proteins from the tobacco
           hornworm, Manduca sexta
    • Abstract: Publication date: Available online 8 January 2015
      Source:Insect Biochemistry and Molecular Biology
      Author(s): Neal T. Dittmer , Guillaume Tetreau , Xiaolong Cao , Haobo Jiang , Ping Wang , Michael R. Kanost
      The insect cuticle is a unique material that covers the exterior of the animal as well as lining the foregut, hindgut, and tracheae. It offers protection from predators and desiccation, defines body shape, and serves as an attachment site for internal organs and muscle. It has demonstrated remarkable variations in hardness, flexibility and elasticity, all the while being light weight, which allows for ease of movement and flight. It is composed primarily of chitin, proteins, catecholamines, and lipids. Proteomic analyses of cuticle from different life stages and species of insects has allowed for a more detailed examination of the protein content and how it relates to cuticle mechanical properties. It is now recognized that several groups of cuticular proteins exist and that they can be classified according to conserved amino acid sequence motifs. We have annotated the genome of the tobacco hornworm, Manduca sexta, for genes that encode putative cuticular proteins that belong to seven different groups: proteins with a Rebers and Riddiford motif (CPR), proteins analogous to peritrophins (CPAP), proteins with a tweedle motif (CPT), proteins with a 44 amino acid motif (CPF), proteins that are CPF-like (CPFL), proteins with an 18 amino acid motif (18 aa), and proteins with two to three copies of a C-X5-C motif (CPCFC). In total we annotated 248 genes, of which 207 belong to the CPR family, the most for any insect genome annotated to date. Additionally, we discovered new members of the CPAP family and determined that orthologous genes are present in other insects. We established orthology between the M. sexta and Bombyx mori genes and identified duplication events that occurred after separation of the two species. Finally, we utilized 52 RNAseq libraries to ascertain gene expression profiles that revealed commonalities and differences between different tissues and developmental stages.
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      PubDate: 2015-01-19T18:28:59Z
       
  • Transcriptomic response of Manduca sexta immune tissues to parasitization
           by the bracovirus associated wasp Cotesia congregata
    • Abstract: Publication date: Available online 10 January 2015
      Source:Insect Biochemistry and Molecular Biology
      Author(s): Germain Chevignon , Sébastien Cambier , Corinne Da Silva , Julie Poulain , Jean-Michel Drezen , Elisabeth Huguet , Sébastien Moreau
      During oviposition, Cotesia congregata parasitoid wasps inject into their host, Manduca sexta, some biological factors such as venom, ovarian fluid and a symbiotic polydnavirus (PDV) named Cotesia congregata bracovirus (CcBV). During parasitism, complex interactions occur between wasp-derived factors and host targets that lead to important modifications in host physiology. In particular, the immune response leading to wasp egg encapsulation is inhibited allowing wasp survival. To date, the regulation of host genes during the interaction had only been studied for a limited number of genes. In this study, we analysed the global impact of parasitism on host gene regulation 24 h post oviposition by high throughput 454 transcriptomic analyses of two tissues known to be involved in the host immune response (hemocytes and fat body). To identify specific effects of parasitism on host transcription at this time point, transcriptomes were obtained from non-treated and parasitized larvae, and also from larvae injected with heat-killed bacteria and double stimulated larvae that were parasitized prior to bacterial challenge. Results showed that, immune challenge by bacteria leads to induction of certain antimicrobial peptide (AMP) genes in M. sexta larvae whether they were parasitized or not prior to bacterial challenge. These results show that at 24 h post oviposition pathways leading to expression of AMP genes are not all inactivated suggesting wasps are in an antiseptic environment. In contrast, at this time point genes involved in phenoloxidase activation and cellular immune responses were globally down-regulated after parasitism in accordance with the observed inhibition of wasp egg encapsulation.
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      PubDate: 2015-01-19T18:28:59Z
       
  • Positive feedback regulation of prothoracicotropic hormone secretion by
           ecdysteroid – A mechanism that determines the timing of
           metamorphosis
    • Abstract: Publication date: Available online 13 January 2015
      Source:Insect Biochemistry and Molecular Biology
      Author(s): Akira Mizoguchi , Manabu Kamimura , Makoto Kiuchi , Hiroshi Kataoka
      When insect larvae have fully grown, prothoracicotropic hormone (PTTH) is released from the brain, triggering the initiation of metamorphic development through stimulation of ecdysteroid secretion by the prothoracic glands. The present study analyzes the mechanism that regulates the occurrence of this PTTH surge. In the silkworm Bombyx mori, the PTTH surge occurs on day 6 of the fifth instar and is preceded by a small rise in hemolymph ecdysteroid titer, which occurs late on day 5. We therefore hypothesized that this rise of ecdysteroid titer is involved in the induction of the PTTH surge. To test this hypothesis, two experiments were conducted. First, a small amount of 20-hydroxyecdysone was injected on day 4, two days before the expected day of the PTTH surge, to simulate the small rise in hemolymph ecdysteroid titer on day 5. This injection led to a precocious surge of PTTH the next day. Next, the hemolymph ecdysteroid titer on day 5 was artificially lowered by injecting ecdysteroid-22-oxidase, which inactivates 20-hydroxyecdysone. After this treatment, the PTTH surge did not occur on day 6 in 80% of the animals. These results indicate that a small rise of the hemolymph ecdysteroid titer plays a critical role in the induction of the PTTH surge. Since basal ecdysteroidogenic activity of the prothoracic glands increases with larval growth, a circulating level of ecdysteroids may convey information about larval maturity to the brain, to coordinate larval growth and metamorphosis. This is the first report in invertebrates to demonstrate positive feedback regulation of the surge of a tropic hormone by a downstream steroid hormone.
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      PubDate: 2015-01-19T18:28:59Z
       
  • Glandular β-glucosidases in juvenile Chrysomelina leaf beetles
           support the evolution of a host-plant-dependent chemical defense
    • Abstract: Publication date: Available online 13 January 2015
      Source:Insect Biochemistry and Molecular Biology
      Author(s): Peter Rahfeld , Wiebke Haeger , Roy Kirsch , Gerhard Pauls , Tobias Becker , Eva Schulze , Natalie Wielsch , Ding Wang , Marco Groth , Wolfgang Brandt , Wilhelm Boland , Antje Burse
      Plant-feeding insects are spread across the entire plant kingdom. Because they chew externally on leaves, leaf beetle of the subtribe Chrysomelina sensu stricto are constantly exposed to life-threatening predators and parasitoids. To counter these pressures, the juveniles repel their enemies by displaying glandular secretions that contain defensive compounds. These repellents can be produced either de novo (iridoids) or by using plant-derived precursors. The autonomous production of iridoids pre-dates the evolution of phytochemical-based defense strategies. Both strategies include hydrolysis of the secreted non-toxic glycosides in the defensive exudates. By combining in vitro as well as in vivo experiments, we show that iridoid de novo producing as well as sequestering species rely on secreted β-glucosidases to cleave the pre-toxins. Our phylogenetic analyses support a common origin of chrysomeline β-glucosidases. The kinetic parameters of these β-glucosidases demonstrated substrate selectivity which reflects the adaptation of Chrysomelina sensu stricto to the chemistry of their hosts during the course of evolution. However, the functional studies also showed that the broad substrate selectivity allows building a chemical defense, which is dependent on the host plant, but does not lead to an “evolutionary dead end”.
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      PubDate: 2015-01-19T18:28:59Z
       
  • Altered tyrosine metabolism and melanization complex formation underlie
           the developmental regulation of melanization in Manduca sexta
    • Abstract: Publication date: Available online 13 January 2015
      Source:Insect Biochemistry and Molecular Biology
      Author(s): Kevin D. Clark
      The study of hemolymph melanization in Lepidoptera has contributed greatly to our understanding of its role in insect immunity. Manduca sexta in particular has been an excellent model for identifying the myriad components of the phenoloxidase (PO) cascade and their activation through exposure to pathogen-associated molecular patterns (PAMPs). However, in a process that is not well characterized or understood, some insect species rapidly melanize upon wounding in the absence of added PAMPs. We sought to better understand this process by measuring wound-induced melanization in four insect species. Of these, only plasma from late 5th instar M. sexta was unable to melanize, even though each contained millimolar levels of the putative melanization substrate tyrosine (Tyr). Analysis of Tyr metabolism using substrate-free plasmas (SFPs) from late 5th instar larvae of each species showed that only M. sexta SFP failed to melanize with added Tyr. In contrast, early instar M. sexta larvae exhibited wound-induced melanization and Tyr metabolism, and SFPs prepared from these larvae melanized in the presence of Tyr. Early instar melanization in M. sexta was associated with the formation of a high mass protein complex that could be observed enzymatically in native gels or by PO-specific immunoblotting. Topical treatment of M. sexta larvae with the juvenile hormone (JH) analog methoprene delayed pupation and increased melanizing ability late in the instar, thus linking development with immunity. Our results demonstrate that melanization rates are highly variable in Lepidoptera, and that developmental stage can be an important factor for melanization within a species. More specifically, we show that the physiological substrate for melanization in M. sexta is Tyr, and that melanization is associated with the formation of a PO-containing protein complex.
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      PubDate: 2015-01-19T18:28:59Z
       
  • Digestive peptidase evolution in holometabolous insects led to a divergent
           group of enzymes in Lepidoptera
    • Abstract: Publication date: Available online 16 January 2015
      Source:Insect Biochemistry and Molecular Biology
      Author(s): Renata O. Dias , Allegra Via , Marcelo M. Brandão , Anna Tramontano , Marcio C. Silva-Filho
      Trypsins and chymotrypsins are well-studied serine peptidases that cleave peptide bonds at the carboxyl side of basic and hydrophobic l-amino acids, respectively. These enzymes are largely responsible for the digestion of proteins. Three primary processes regulate the activity of these peptidases: secretion, precursor (zymogen) activation and substrate-binding site recognition. Here, we present a detailed phylogenetic analysis of trypsins and chymotrypsins in three orders of holometabolous insects and reveal divergent characteristics of Lepidoptera enzymes in comparison with those of Coleoptera and Diptera. In particular, trypsin subsite S1 was more hydrophilic in Lepidoptera than in Coleoptera and Diptera, whereas subsites S2–S4 were more hydrophobic, suggesting different substrate preferences. Furthermore, Lepidoptera displayed a lineage-specific trypsin group belonging only to the Noctuidae family. Evidence for facilitated trypsin auto-activation events were also observed in all the insect orders studied, with the characteristic zymogen activation motif complementary to the trypsin active site. In contrast, insect chymotrypsins did not seem to have a peculiar evolutionary history with respect to their mammal counterparts. Overall, our findings suggest that the need for fast digestion allowed holometabolous insects to evolve divergent groups of peptidases with high auto-activation rates, and highlight that the evolution of trypsins led to a most diverse group of enzymes in Lepidoptera.
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      PubDate: 2015-01-19T18:28:59Z
       
  • GC/MS-based metabolomic studies reveal key roles of glycine
           in regulating silk synthesis in silkworm, Bombyx mori
    • Abstract: Publication date: February 2015
      Source:Insect Biochemistry and Molecular Biology, Volume 57
      Author(s): Quanmei Chen , Xinyu Liu , Ping Zhao , Yanhui Sun , Xinjie Zhao , Ying Xiong , Guowang Xu , Qingyou Xia
      Metabolic profiling of silkworm, especially the factors that affect silk synthesis at the metabolic level, is little known. Herein, metabolomic method based on gas chromatography-mass spectrometry was applied to identify key metabolic changes in silk synthesis deficient silkworms. Forty-six differential metabolites were identified in Nd group with the defect of silk synthesis. Significant changes in the levels of glycine and uric acid (up-regulation), carbohydrates and free fatty acids (down-regulation) were observed. The further metabolomics of silk synthesis deficient silkworms by decreasing silk proteins synthesis using knocking out fibroin heavy chain gene or extirpating silk glands operation showed that the changes of the metabolites were almost consistent with those of the Nd group. Furthermore, the increased silk yields by supplying more glycine or its related metabolite confirmed that glycine is a key metabolite to regulate silk synthesis. These findings provide important insights into the regulation between metabolic profiling and silk synthesis.
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      PubDate: 2015-01-19T18:28:59Z
       
  • Molecular characterization and functional expression of the Apis mellifera
           voltage-dependent Ca2+ channels
    • Abstract: Publication date: Available online 17 January 2015
      Source:Insect Biochemistry and Molecular Biology
      Author(s): Thierry Cens , Matthieu Rousset , Claude Collet , Mercedes Charreton , Lionel Garnery , Yves Le Conte , Mohamed Chahine , Jean-Christophe Sandoz , Pierre Charnet
      Voltage-gated Ca2+ channels allow the influx of Ca2+ ions from the extracellular space upon membrane depolarization and thus serve as a transducer between membrane potential and cellular events initiated by Ca2+ transients. Most insects are predicted to possess three genes encoding Cavα, the main subunit of Ca2+ channels, and several genes encoding the two auxiliary subunits, Cavβ and Cavα2δ; however very few of these genes have been cloned so far. Here, we cloned three full-length cDNAs encoding the three Cavα subunits (AmelCav1a, AmelCav2a and AmelCav3a), a cDNA encoding a novel variant of the Cavβ subunit (AmelCavβc), and three full-length cDNAs encoding three Cavα2δ subunits (AmelCavα2δ1 to 3) of the honeybee Apis mellifera. We identified several alternative or mutually exclusive exons in the sequence of the AmelCav2 and AmelCav3 genes. Moreover, we detected a stretch of glutamine residues in the C-terminus of the AmelCav1 subunit that is reminiscent of the motif found in the human Cav2.1 subunit of patients with Spinocerebellar Ataxia type 6. All these subunits contain structural domains that have been identified as functionally important in their mammalian homologues. For the first time, we could express three insect Cavα subunits in Xenopus oocytes and we show that AmelCav1a, 2a and 3a form Ca2+ channels with distinctive properties. Notably, the co-expression of AmelCav1a or AmelCav2a with AmelCavβc and AmCavα2δ1 produces High Voltage-Activated Ca2+ channels. On the other hand, expression of AmelCav3a alone leads to Low Voltage-Activated Ca2+ channels.
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      PubDate: 2015-01-19T18:28:59Z
       
  • Regulation of histone H3 phosphorylation at serine 10 in PTTH-stimulated
           prothoracic glands of the silkworm, Bombyx mori
    • Abstract: Publication date: February 2015
      Source:Insect Biochemistry and Molecular Biology, Volume 57
      Author(s): Shi-Hong Gu , Yun-Chin Hsieh
      A complex signaling network appears to be involved in prothoracicotropic hormone (PTTH)-stimulated ecdysteroidogenesis in insect prothoracic glands (PGs). In the present study, we investigated the localization of phosphorylated extracellular signal-regulated kinase (ERK) in PTTH-stimulated PGs in Bombyx mori. The nuclear effect of PTTH was further studied by examining phosphorylation of histone H3 at serine 10. Results showed that in PTTH-stimulated PGs, higher phosphorylated ERK was detected in nuclear fraction compared to that in cytosolic fraction. PTTH treatment in vitro appears to rapidly enhance the transcriptional activation-associated histone H3 phosphorylation at serine 10. PTTH stimulated histone H3 phosphorylation in a time-dependent manner. Injection of PTTH into day-6 last instar larvae greatly increased histone H3 phosphorylation, verifying the in vitro effect. The stimulation of histone H3 phosphorylation by PTTH appears to be developmentally regulated. PTTH-stimulated histone H3 phosphorylation was greatly reduced in Ca2+-free saline or by pretreatment with a potent and specific inhibitor of phospholipase C (PLC), U73122. When PGs were treated with agents that directly elevate the intracellular Ca2+ concentration (either A23187 or thapsigargin), a greatly increase in histone H3 phosphorylation at serine 10 was observed, indicating Ca2+-dependency of histone H3 phosphorylation stimulated by PTTH. In addition, PTTH-stimulated histone H3 phosphorylation was partially reduced by U0126, a specific mitogen-activated protein kinase (MAPK)/ERK kinase (MEK) inhibitor, indicating the involvement of ERK. However, pretreatment with LY294002, a phosphoinositide 3-kinase (PI3K) inhibitor, did not inhibit PTTH-stimulated histone H3 phosphorylation, implying that PI3K signaling is not related to PTTH-stimulated histone H3 phosphorylation. Taken together, these results suggest that PTTH-stimulated histone H3 phosphorylation at serine 10 is mediated by Ca2+/ERK signaling in B. mori PGs.
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      PubDate: 2015-01-19T18:28:59Z
       
  • TIL-type protease inhibitors may be used as targeted resistance factors to
           enhance silkworm defenses against invasive fungi
    • Abstract: Publication date: February 2015
      Source:Insect Biochemistry and Molecular Biology, Volume 57
      Author(s): Youshan Li , Ping Zhao , Huawei Liu , Xiaomeng Guo , Huawei He , Rui Zhu , Zhonghuai Xiang , Qingyou Xia
      Entomopathogenic fungi penetrate the insect cuticle using their abundant hydrolases. These hydrolases, which include cuticle-degrading proteases and chitinases, are important virulence factors. Our recent findings suggest that many serine protease inhibitors, especially TIL-type protease inhibitors, are involved in insect resistance to pathogenic microorganisms. To clarify the molecular mechanism underlying this resistance to entomopathogenic fungi and identify novel genes to improve the silkworm antifungal capacity, we conducted an in-depth study of serine protease inhibitors. Here, we cloned and expressed a novel silkworm TIL-type protease inhibitor, BmSPI39. In activity assays, BmSPI39 potently inhibited the virulence protease CDEP-1 of Beauveria bassiana, suggesting that it might suppress the fungal penetration of the silkworm integument by inhibiting the cuticle-degrading proteases secreted by the fungus. Phenol oxidase activation studies showed that melanization is involved in the insect immune response to fungal invasion, and that fungus-induced excessive melanization is suppressed by BmSPI39 by inhibiting the fungal cuticle-degrading proteases. To better understand the mechanism involved in the inhibition of fungal virulence by protease inhibitors, their effects on the germination of B. bassiana conidia was examined. BmSPI38 and BmSPI39 significantly inhibited the germination of B. bassiana conidia. Survival assays showed that BmSPI38 and BmSPI39 markedly improved the survival rates of silkworms, and can therefore be used as targeted resistance proteins in the silkworm. These results provided new insight into the molecular mechanisms whereby insect protease inhibitors confer resistance against entomopathogenic fungi, suggesting their potential application in medicinal or agricultural fields.
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      PubDate: 2015-01-19T18:28:59Z
       
  • Allatostatin-C reversibly blocks the transport of citrate out of the
           mitochondria and inhibits juvenile hormone synthesis in mosquitoes
    • Abstract: Publication date: February 2015
      Source:Insect Biochemistry and Molecular Biology, Volume 57
      Author(s): Marcela Nouzova , Crisalejandra Rivera-Perez , Fernando G. Noriega
      Aedes aegypti allatostatin-C (AeaAST-C or PISCF-AST) is a strong and fast reversible inhibitor of juvenile hormone III (JH III) synthesis by the corpora allata (CA) of mosquitoes; however, its mechanism of action remains poorly understood. AeaAST-C showed no inhibitory activity in the presence of any of the intermediate precursors of JH III indicating that the AeaAST-C target is located before the entry of acetyl-CoA in the pathway. Stimulation experiments using different sources of carbon (glucose, pyruvate, acetate and citrate) suggest that AST-C acts after pyruvate is transformed to citrate in the mitochondria. In vitro inhibition of the citrate mitochondrial carrier (CIC) mimicked the effect of AeaAST-C, and was overridden by addition of citrate or acetate. Our results provide compelling evidence that AeaAST-C inhibits JH III synthesis by blocking the CIC carrier that transports citrate from the mitochondria to the cytosol, obstructing the production of cytoplasmic acetyl-CoA that sustains JH III synthesis in the CA of mosquitoes.
      Graphical abstract image

      PubDate: 2015-01-19T18:28:59Z
       
  • The structural sheath protein of aphids is required for phloem feeding
    • Abstract: Publication date: Available online 17 December 2014
      Source:Insect Biochemistry and Molecular Biology
      Author(s): Torsten Will , Andreas Vilcinskas
      Aphids produce two types of saliva that mediate their interactions with plants. Watery saliva is secreted during cell penetration and ingestion, whereas gel saliva is secreted during stylet movement through the apoplast where it forms a sheath around the stylet to facilitate penetration and seal puncture sites on cell membranes. In order to study the function of the sheath when aphids interact with plants, we used RNA interference (RNAi) to silence the aphid structural sheath protein (SHP) in the pea aphid Acyrthosiphon pisum. The injection of 50 ng of double stranded RNA completely disrupted sheath formation, as confirmed by scanning electron microscopy. Aphid behavior was monitored using the electrical penetration graph technique, revealing that disrupted sheath formation prevented efficient long-term feeding from sieve tubes, with a silencing effect on reproduction but not survival. We propose that sealing the stylet penetration site in the sieve tube plasma membrane is part of a two-step mechanism to suppress sieve-tube occlusion by preventing calcium influx into the sieve tube lumen. The SHP is present in several aphid species and silencing has a similar impact to aphid-resistant plants, suggesting that SHP is an excellent target for RNAi-mediated pest control.
      Graphical abstract image

      PubDate: 2014-12-19T06:47:25Z
       
  • An atypical residue in the pore of Varroa destructor GABA-activated RDL
           receptors affects picrotoxin block and thymol modulation
    • Abstract: Publication date: December 2014
      Source:Insect Biochemistry and Molecular Biology, Volume 55
      Author(s): Kerry L. Price , Sarah C.R. Lummis
      GABA-activated RDL receptors are the insect equivalent of mammalian GABAA receptors, and play a vital role in neurotransmission and insecticide action. Here we clone the pore lining M2 region of the Varroa mite RDL receptor and show that it has 4 atypical residues when compared to M2 regions of most other insects, including bees, which are the major host of Varroa mites. We create mutant Drosophila RDL receptors containing these substitutions and characterise their effects on function. Using two electrode voltage clamp electrophysiology we show that one substitution (T6′M) ablates picrotoxin inhibition and increases the potency of GABA. This mutation also alters the effect of thymol, which enhances both insect and mammalian GABA responses, and is widely used as a miticide. Thymol decreases the GABA EC50 of WT receptors, enhancing responses, but in T6′M-containing receptors it is inhibitory. The other 3 atypical residues have no major effects on either the GABA EC50, the picrotoxin potency or the effect of thymol. In conclusion we show that the RDL 6′ residue is important for channel block, activation and modulation, and understanding its function also has the potential to prove useful in the design of Varroa-specific insecticidal agents.
      Graphical abstract image

      PubDate: 2014-12-15T09:05:32Z
       
 
 
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