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

Clark, Kevin D.; Strand, Michael R.

doi:10.1074/jbc.m113.459222

Cited by 60 publications

(52 citation statements)

References 54 publications

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“…By identifying 66 proteins (10 -80 kDa) in gel slices 1-3 (80 to Ͼ200 kDa) (Table S5), we validated and extended the previous observations of such complexes (16,19,43,45,46). Nearly half (32) of them (e.g.…”

Section: Table II Distribution Of Selected Immunity-related Proteins supporting

confidence: 66%

Changes in the Plasma Proteome of Manduca sexta Larvae in Relation to the Transcriptome Variations after an Immune Challenge: Evidence for High Molecular Weight Immune Complex Formation

Cao

Zhang

et al. 2016

Molecular & Cellular Proteomics

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Manduca sexta is a lepidopteran model widely used to study insect physiological processes, including innate immunity. In this study, we explored the proteomes of cellfree hemolymph from larvae injected with a sterile buffer (C for control) or a mixture of bacteria (I for induced). Of the 654 proteins identified, 70 showed 1.67 to >200-fold abundance increases after the immune challenge; 51 decreased to 0 -60% of the control levels. While there was no strong parallel between plasma protein levels and their transcript levels in hemocytes or fat body, the mRNA level changes (i.e. I/C ratios of normalized read numbers) in the tissues concurred with their protein level changes (i.e. I/C ratios of normalized spectral counts) with correlation coefficients of 0.44 and 0.57, respectively. Better correlations support that fat body contributes a more significant portion of the plasma proteins involved in various aspects of innate immunity. Consistently, ratios of mRNA and protein levels were better correlated for immunity-related proteins than unrelated ones. There is a set of proteins whose apparent molecular masses differ considerably from the calculated M r 's, suggestive of posttranslational modifications. In addition, some low M r proteins were detected in the range of 80 to >300 kDa on a reducing SDS-polyacrylamide gel, indicating the existence of high M r covalent complexes. We identified 30 serine proteases and their homologs, 11 of which are known members of an extracellular immune signaling network. Along with our quantitative transcriptome data, the protein identification, inducibility, and association provide leads toward a focused exploration of humoral immunity in M. sexta. Molecular & Cellular

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Section: Table II Distribution Of Selected Immunity-related Proteins supporting

confidence: 66%

Changes in the Plasma Proteome of Manduca sexta Larvae in Relation to the Transcriptome Variations after an Immune Challenge: Evidence for High Molecular Weight Immune Complex Formation

Cao

Zhang

et al. 2016

Molecular & Cellular Proteomics

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“…The endogenous pattern recognition proteins and proHP1 in plasma apparently do not initiate the system under the normal physiological condition. Perhaps, spontaneous oligomerization of these system components can trigger melanization and additional association of the oligomers on microbial surfaces enhance the potency and locality of this response (Yu et al, 2003; Clark and Strand, 2013; He et al, 2016). …”

Section: Resultsmentioning

confidence: 99%

In search of a function of Manduca sexta hemolymph protease-1 in the innate immune system

Yang

Wang

et al. 2016

Insect Biochemistry and Molecular Biology

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Extracellular serine protease cascades mediate immune signaling and responses in insects. In the tobacco hornworm Manduca sexta, nearly 30 serine proteases (SPs) and their homologs SPHs) are cloned from hemocytes and fat body. Some of them participate in prophenoloxidase proPO) activation and proSpätzle processing. Here we report the cDNA cloning of hemolymph protease-1b (HP1b), which is 90% identical and 95% similar to HP1a (formerly HP1). The HP1a and HP1b mRNA levels in hemocytes was down- and up-regulated after an immune challenge, respectively. Quantitative real-time polymerase chain reactions revealed their tissue-specific and development-dependent expression, mostly in hemocytes of the feeding larvae. We isolated HP1 precursor (proHP1) from larval hemolymph and observed micro-heterogeneity caused by N-linked glycosylation. Supplementation of the purified proHP1 to plasma samples from naïve larvae or induced ones injected with bacteria caused a small PO activity increase, much lower than those elicited by recombinant proHP1a/b, but no proteolytic cleavage was detected in the zymogens. Incubation of proHP1a/b or their catalytic domains with a cationic detergent, cetylpyridinium chloride, induced an amidase activity that hydrolyzed LDLH-p-nitroanilide. Since LDLH corresponds to the P4–P1 region before the proteolytic activation site of proHP6, we propose that the active but uncleaved proHP1 may cut proHP6 to generate HP6 that in turn activates proPAP1 and proHP8. The catalytic domain of HP1a/b, which by itself does not activate purified proHP6 or hydrolyze LDLH-p-nitroanilide, somehow generated active HP6, HP8, PAP1 and PO in plasma. Together, these results indicate that proHP1 participates in the proPO activation system, although detailed mechanism needs further exploration.

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“…Therefore, it is plausible that B. mori C-type lectins play important roles not only in the recognition of invading pathogens, but also in the initiation of proPO activation by means of BmSPH1 and BmSPH2. Clark and Strand (2013) reported that SPH1 and a kind of C-type lectin (C-type lectin 21) formed a complex with PO in B. mori (Clark and Strand, 2013). Thus, protein complexes consisting of BmSPH1, BmSPH2 and B. mori C-type lectins observed in our examination (Fig.…”

Section: Bmsphs In Plasma Are Concentrated In Nodules By B Mori Ctypmentioning

confidence: 47%

“…Thus, protein complexes consisting of BmSPH1, BmSPH2 and B. mori C-type lectins observed in our examination (Fig. 3) might be precursors of the PO-containing complex observed by Clark and Strand (2013). It is possible that protein complexes consisting of BmSPH1, BmSPH2 and B. mori C-type lectins help PO to congregate around the surface of pathogens, resulting in the melanin deposition on the pathogens.…”

Section: Bmsphs In Plasma Are Concentrated In Nodules By B Mori Ctypmentioning

confidence: 63%

Mechanisms of nodule-specific melanization in the hemocoel of the silkworm, Bombyx mori

Shi¹,

Mang²,

Fu³

et al. 2016

Insect Biochemistry and Molecular Biology

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

Cited by 60 publications

References 54 publications

Changes in the Plasma Proteome of Manduca sexta Larvae in Relation to the Transcriptome Variations after an Immune Challenge: Evidence for High Molecular Weight Immune Complex Formation

Changes in the Plasma Proteome of Manduca sexta Larvae in Relation to the Transcriptome Variations after an Immune Challenge: Evidence for High Molecular Weight Immune Complex Formation

In search of a function of Manduca sexta hemolymph protease-1 in the innate immune system

Mechanisms of nodule-specific melanization in the hemocoel of the silkworm, Bombyx mori

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