Identification by subtractive suppression hybridization of bacteria-induced genes expressed in Manduca sexta fat body

Zhu, Yifei; Johnson, Timothy J.; Myers, Alisoncowieandrichardm .; Kanost, Michael R.

doi:10.1016/s0965-1748(03)00028-6

Cited by 110 publications

(96 citation statements)

References 115 publications

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“…In addition, we never detected tyramine or DA in B. mori plasma in the first 20 min after collection from a wound site. Thus, although Tyr decarboxylase and DOPA decarboxylase have both been suggested to play roles in converting Tyr to tyramine and DOPA to DA for use by POs (49,(53)(54)(55), we see no evidence for this during the rapid melanization of B. mori plasma from a wound site.…”

Section: Discussioncontrasting

confidence: 61%

Hemolymph Melanization in the Silkmoth Bombyx mori Involves Formation of a High Molecular Mass Complex That Metabolizes Tyrosine

Clark

Strand

2013

Journal of Biological Chemistry

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Background:The phenoloxidase (PO) cascade regulates the melanization of blood in insects and other arthropods. Results: Hemolymph from Bombyx mori rapidly melanizes after wounding and forms a complex that uses tyrosine as the substrate. Conclusion: Tyrosine usage requires formation of a phenoloxidase-containing complex. Significance: Processing of prophenoloxidase is an intermediate step in the phenoloxidase cascade.

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Section: Discussioncontrasting

confidence: 61%

Hemolymph Melanization in the Silkmoth Bombyx mori Involves Formation of a High Molecular Mass Complex That Metabolizes Tyrosine

Clark

Strand

2013

Journal of Biological Chemistry

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“…In vivo RNA interference (RNAi) 3 mediated knock-down of Noduler especially affected the nodulation response thus suggesting a role for this protein in immunity. Similar proteins have been reported earlier from other lepidopterans but their role in immunity was not understood (15,16).…”

supporting

confidence: 74%

“…1B). Noduler homologues have been reported to be immune up-regulated in other lepidopterans (one homologue identified in each of Samia cynthia ricini, Lonomia obliqua, Bombyx mori, Manduca sexta and Hyphantria cunea) (15,16,22,23). Among the other insects, three Noduler homologues were found in Anopheles gambiae, four in Drosophila melanogaster, two in D. pseudoobscura, three in Aedes aegypti, three in Tribolium castaneum, and one in Apis mellifera (24,25).…”

Section: A Reeler Domain Spans a Major Part Of Noduler Proteinmentioning

confidence: 99%

Noduler, A Novel Immune Up-Regulated Protein Mediates Nodulation Response in Insects

Gandhe

John

Nagaraju

2007

The Journal of Immunology

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Insect immune system comprises of both humoral and cellular defenses. Nodulation is one of the major, yet very poorly understood cellular responses against microbial infections in insects. Through screening for novel immune genes from an Indian saturniid silkmoth Antheraea mylitta, we identified a protein up-regulated in hemolymph within minutes upon bacterial challenge. We have shown here, for first time, the involvement of this novel protein in mediating nodulation response against bacteria and hence designated it as Noduler. Noduler possessed a characteristic reeler domain found in several extracellular matrix vertebrate proteins. Noduler was shown in vitro to bind a wide range of bacteria, yeast, and also insect hemocytes. Furthermore, Noduler specifically bound LPS, lipotechoic acid, and β-1, 3 glucan components of microbial cell walls. RNA-interference mediated knock-down of the Noduler resulted in significant reduction in the number of nodules and consequent increase in bacterial load in larval hemolymph. The results suggest that the Noduler is widely conserved and is involved in very early clearance of bacteria by forming nodules of hemocytes and bacterial complexes in insects. The results would promote further studies for understanding of the crucial but hitherto overlooked nodulation mechanism in insects and also provide cues for the study of similar mammalian proteins whose function is not understood.

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“…A plasmid containing a 902-bp serpin-4 cDNA fragment (GenBank TM accession number BG835792) was obtained from Dr. Hugh M. Robertson, University of Illinois. A 286-bp serpin-5 cDNA fragment was amplified by PCR from clone N27 (BI262723) (31). The purified cDNA fragments were labeled with [␣-sequenced using ABI PRISM BigDye Terminator Cycle Sequencing Ready Reaction kit (PerkinElmer Life Sciences).…”

Section: Methodsmentioning

confidence: 99%

Manduca sexta Serpin-4 and Serpin-5 Inhibit the Prophenol Oxidase Activation Pathway

Tong

Kanost

2005

Journal of Biological Chemistry

108

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Infection stimulates the innate immune responses of insects, including activation of prophenol oxidase (pro-PO) in plasma as the last step of a serine protease cascade. To investigate the roles of protease inhibitors in regulating this pathway, we cloned cDNAs for two new serpins (serpin-4 and serpin-5) from the tobacco hornworm, Manduca sexta. Serpin-4 and serpin-5 mRNAs are constitutively expressed at a low level in larval hemocytes and fat body and increased dramatically upon bacterial challenge. These serpins are present in larval plasma at ϳ3 (serpin-4) and ϳ1 g/ml (serpin-5) and increased 3-8-fold by 24 h after injection of bacteria or fungi. Recombinant serpin-4 and serpin-5 decreased pro-PO activation when added to plasma, but they did not directly inhibit the pro-PO-activating proteases. Instead, they apparently regulate the pathway by inhibiting one or more target proteases upstream of the pro-PO-activating proteases.Prophenol oxidase (pro- PO) 1 activation is an innate immune response in arthropods. Upon injury or infection, a pro-PO zymogen in plasma is activated by a specific protease. Phenol oxidase (PO) hydroxylates monophenols to o-diphenols and then oxidizes o-diphenols to quinones, which can polymerize to form melanin at the injury site or around invading organisms (1, 2). Quinones may also be involved in the production of cytotoxic molecules such as superoxides and hydroxyl radicals that could help kill the invading organisms (1, 3).A serine protease cascade functions in pro-PO activation (2, 4, 5). The final protease in the pathway, pro-PO-activating protease (PAP) (also called pro-PO-activating enzyme or factor) has been identified from several insects, including the tobacco hornworm, Manduca sexta (6 -8), the silkworm, Bombyx mori (9), a beetle, Holotrichia diomphalia (10, 11), and from a crustacean, the crayfish Pacifastacus leniusculus (12). These PAPs contain one or two clip domains at their amino terminus and a serine protease domain at their carboxyl terminus (13). PAPs are synthesized as zymogens, which are activated by a specific proteolytic cleavage between the clip domain and the protease domain. However, protease(s) that activate PAPs have not yet been characterized.Serpins present in plasma regulate the pro-PO activation pathway (4,5,(13)(14)(15)(16). Serpins are serine protease inhibitors, about 400 amino acid residues long (17-20), with an exposed reactive center loop near their carboxyl terminus (21,22). The P1 residue located in this loop determines the primary specificity of inhibition. Four serpins from M. sexta have been characterized previously. The serpin-1 gene encodes 12 variants with different inhibitory selectivity, due to alternative splicing of exons that encode the reactive center loop (23-25). Serpin-1 mRNA is constitutively expressed in the larval fat body and at a lower level in hemocytes (26). One of the reactive site variants, serpin-1J, inhibits pro-PO activation in plasma by inhibiting PAPs (8, 25). Serpin-2 is an intracellular protein (27), expressed in ...

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Identification by subtractive suppression hybridization of bacteria-induced genes expressed in Manduca sexta fat body

Cited by 110 publications

References 115 publications

Hemolymph Melanization in the Silkmoth Bombyx mori Involves Formation of a High Molecular Mass Complex That Metabolizes Tyrosine

Hemolymph Melanization in the Silkmoth Bombyx mori Involves Formation of a High Molecular Mass Complex That Metabolizes Tyrosine

Noduler, A Novel Immune Up-Regulated Protein Mediates Nodulation Response in Insects

Manduca sexta Serpin-4 and Serpin-5 Inhibit the Prophenol Oxidase Activation Pathway

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