Immunodominant role of CCHA subunit of Concholepas hemocyanin is associated with unique biochemical properties

Becker, María; Fuentes, Alejandra; Campo, Miguel del; Manubens, Augusto; Nova, Esteban; Oliva, Harold; Faunes, Fernando; Valenzuela, Manuel; Campos‐Vallette, Marcelo; Aliaga, Álvaro; Ferreira, Jorge; Ioannes, Alfredo E. De; Ioannes, Pablo De; Moltedo, Bruno

doi:10.1016/j.intimp.2008.12.011

Cited by 30 publications

(23 citation statements)

References 59 publications

(73 reference statements)

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“…Many authors have proposed different factors to explain these properties, such as the large size, the complex quaternary structure with D5-like symmetry, the xenogenicity, and the sugar moiety content of hemocyanins. However, our studies evaluating the immunosti-mulatory properties of the isolated subunits of CCH have shown that the large size and the complex quaternary structure do not determine the immunogenicity and antitumor effects of hemo-cyanins in a mouse model of SBC [12]. This observation has been supported by another report that assessed the immunogenicity of subunits isolated from R. thomasiana hemocyanin [13].…”

Section: Introductionmentioning

confidence: 59%

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Enhanced structural stability of Concholepas hemocyanin increases its immunogenicity and maintains its non‐specific immunostimulatory effects

et al. 2012

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Hemocyanins, which boost the immune system of mammals, have been used as carrieradjuvants to promote Ab production against haptens and peptides, as immunostimulants during therapy for bladder carcinoma and as a component in therapeutic vaccines for cancer. These biomedical applications have led to growing interest in obtaining hemocyanins with high immunogenicity. Here, we study the immunological properties of a modified oxidized Concholepas concholepas hemocyanin (Ox-CCH) obtained by the oxidation of its carbohydrates using sodium periodate. We assessed the internalization of Ox-CCH into DCs and its immunogenicity and antitumor effects. Transmission electron microscopy showed no changes in Ox-CCH quaternary structure with respect to native CCH, although proteolytic treatment followed by SDS-PAGE analysis demonstrated that Schiff bases were formed. Interestingly, DCs internalized Ox-CCH faster than CCH, mainly through macropinocytosis. During this process, Ox-CCH remained inside endosome-like structures for a longer period. Mouse immunization experiments demonstrated that Ox-CCH is more immunogenic and a better carrier than CCH. Moreover, Ox-CCH showed a significant antitumor effect in the B16F10 melanoma model similar to that produced by CCH, inducing IFN-γ secretion. Together, these data demonstrate that the aldehydes formed by the periodate oxidation of sugar moieties stabilizes the CCH structure, increasing its adjuvant/immunostimulatory carrier effects.

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Section: Introductionmentioning

confidence: 59%

“…CCH subunits (CCHA and CCHB) were purified according to the methods described by Becker et al [12]. All solutions containing hemocyanins and buffers for their dialysis were maintained at 4…”

Section: Hemocyaninsmentioning

confidence: 99%

Enhanced structural stability of Concholepas hemocyanin increases its immunogenicity and maintains its non‐specific immunostimulatory effects

et al. 2012

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“…In particular, our previous research found that human IgG not only contained several conserved regions with shrimp hemocyanin, but could also cross-react with rabbit anti-shrimp hemocyanin antibodies [13], [14]. In addition, hemocyanin in mollusks and arthropods has been documented to possess several immune functions, such as phenoloxidase [15], antiviral [16], antibacterial [17], antitumoral [18], hemolytic [19] and hemagglutinative [14] activities. Therefore, it is of great interest to further investigate whether human IgG also possesses some antigen nonspecific immune properties.…”

Section: Introductionmentioning

confidence: 99%

Characterization of a Novel Hemolytic Activity of Human IgG Fractions Arising from Diversity in Protein and Oligosaccharide Components

et al. 2014

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Human IgG is a well-established multifunctional antigen specific immunoglobulin molecule of the adaptive immune system. However, an antigen nonspecific immunological function of human IgG has never been reported. In this study, human IgG was isolated using ammonium sulfate fractional precipitation and diethylaminoethanol (DEAE) cellulose 52 ion exchange chromatography, from which h-IgG and hs-IgG fractions were purified on the basis of their differential binding to rabbit anti-shrimp hemocyanin antibody (h) and rabbit anti-shrimp hemocyanin's small subunit antibody (hs), respectively. We found that h-IgG had a higher hemolytic activity than hs-IgG against erythrocytes from humans, rabbits, mice and chickens, whereas the control IgG showed negligible activity. h-IgG could interact directly with erythrocyte membranes, and this interaction was suppressed by high molecular weight osmoprotectants, showing that it may follow a colloid-osmotic mechanism. In comparative proteomics and glycomics studies, h-IgG and hs-IgG yielded 20 and 5 significantly altered protein spots, respectively, on a 2-D gel. The mean carbohydrate content of h-IgG and hs-IgG was approximately 3.6- and 2-fold higher than that of IgG, respectively, and the α-d-mannose/α-d-glucose content was in the order of h-IgG>hs-IgG>IgG. In this study, a novel antigen nonspecific immune property of human IgG was investigated, and the diversity in the protein constituents and glycosylation levels may have functional signficance.

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“…By contrast, in C. concholepas, which contains two subunits, the decamers are formed by two types of intermingled subunits that form a heterodecamer. Consequently, their association in pairs results in heterodidecamers (De Ioannes et al 2004;Becker et al 2009). Thus, the structural differences between these hemocyanins raise problems that remain to be solved with regard to their synthesis.…”

Section: Introductionmentioning

confidence: 99%

“…KLH, CCH, and several other gastropod hemocyanins are under extensive study in order to determine their biochemical and immune properties (Moltedo et al 2006;Siddiqui et al 2007;Dolashka-Angelova et al 2007;Tchorbanov et al 2008;Idakieva et al 2008;Becker et al 2009;Jaenicke et al 2010), to resolve their structure (Hartmann et al 2004;Gatsogiannis and Markl 2009), and to identify their genes (Miller et al 1998;Lieb et al 2001Lieb et al , 2004Altenhein et al 2002;Bergmann et al 2006Bergmann et al , 2007. The biosynthesis of hemocyanins in molluscs has been studied previously (Ruth et al 1988(Ruth et al , 2000van Holde and Miller 1995;Taylor and Anstiss 1999;Albrecht et al 2001;Streit et al 2005).…”

Section: Introductionmentioning

confidence: 99%

Concholepas hemocyanin biosynthesis takes place in the hepatopancreas, with hemocytes being involved in its metabolism

et al. 2010

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Hemocyanins are copper-containing glycoproteins in some molluscs and arthropods, and their best-known function is O(2) transport. We studied the site of their biosynthesis in the gastropod Concholepas concholepas by using immunological and molecular genetic approaches. We performed immunohistochemical staining of various organs, including the mantle, branchia, and hepatopancreas, and detected C. concholepas hemocyanin (CCH) molecules in circulating and tissue-associated hemocytes by electron microscopy. To characterize the hemocytes, we purified them from hemolymph. We identified three types of granular cells. The most abundant type was a phagocyte-like cell with small cytoplasmic granules. The second type contained large electron-dense granules. The third type had vacuoles containing hemocyanin molecules suggesting that synthesis or catabolism occurred inside these cells. Our failure to detect cch-mRNA in hemocytes by reverse transcription with the polymerase chain reaction (RT-PCR) led us to propose that hemocytes instead played a role in CCH metabolism. This hypothesis was supported by colloidal gold staining showing hemocyanin molecules in electron-dense granules inside hemocytes. RT-PCR analysis, complemented by in situ hybridization analyses with single-stranded antisense RNAs as specific probes, demonstrated the presence of cch-mRNA in the hepatopancreas; this was consistent with the specific hybridization signal and confirmed the hepatopancreas as the site of CCH synthesis. Finally, we investigated the possibility that CCH catabolism in hemocytes was involved in the host immune response and in the generation of secondary metabolites such as antimicrobial peptides and phenoloxidase.

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Immunodominant role of CCHA subunit of Concholepas hemocyanin is associated with unique biochemical properties

Cited by 30 publications

References 59 publications

Enhanced structural stability of Concholepas hemocyanin increases its immunogenicity and maintains its non‐specific immunostimulatory effects

Enhanced structural stability of Concholepas hemocyanin increases its immunogenicity and maintains its non‐specific immunostimulatory effects

Characterization of a Novel Hemolytic Activity of Human IgG Fractions Arising from Diversity in Protein and Oligosaccharide Components

Concholepas hemocyanin biosynthesis takes place in the hepatopancreas, with hemocytes being involved in its metabolism

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