Fabrizio Ceciliani

The systemic reaction during inflammation: the acute-phase proteins.

The acute-phase response consists in a large number of behavioural, physiologic, biochemical, and nutritional changes involving many organ systems distant from the site, or sites, of inflammation. One of the most investigated, but still not well understood, characteristic of the acute phase is the up-regulation, or down- regulation, of many plasma proteins, known as the acute-phase proteins. The changes in the concentrations of these positive acute-phase proteins and negative acute-phase proteins are due to changes in their liver production. Their increase may vary from 25 percent to 1000 fold, as in the case of C-reactive protein and serum amyloid A. This review summarises the recent advances that have been acquired on the acute-phase proteins, in particular their function in pathologies such as infections or inflammatory lesions.

Acute phase proteins in ruminants

The physiological response to infections and injuries involves local inflammation and the initiation of events leading to a systemic response, also called acute phase reaction (APR). This multiplicity of changes is distant from the site of injury, and includes fever, leukocytosis and quantitative and qualitative modification of a group of non-structurally related proteins present in blood and other biological fluids, collectively named Acute Phase Proteins (APP). Proteomic investigations of serum or plasma following natural or experimental infection frequently reveal substantial alterations in the APP, several of which are high abundance proteins in these fluids. The present review will focus on the results of recent research on ruminant APP. Highlight points will include: - The structure and the functions of the main APPs in ruminants, as well as the regulatory mechanisms that trigger their systemic and local expression in both physiological and pathological conditions.- The clinical aspects of APPs in ruminants, including the current and future application to veterinary diagnosis and animal production.- The APP in small and wildlife ruminants.- Alteration in APP detected by proteomic investigations.

The Major Surface Protein of Wolbachia Endosymbionts in Filarial Nematodes Elicits Immune Responses through TLR2 and TLR4

More than 150 million humans in tropical countries are infected by filarial nematodes which harbor intracellular bacterial endosymbionts of the genus Wolbachia (Rickettsiales). These bacteria have been implicated in adverse effects of drug treatment in filariasis. The present study provides evidence that purified major Wolbachia surface protein (rWSP) acts as an inducer of the innate immune system through TLR2 and TLR4: 1) recombinant, stringently purified rWSP elicited the release of TNF-α, IL-12, and IL-8 from cultured blood cells of both Onchocerca volvulus-infected and uninfected people; 2) the inflammatory response to rWSP challenge was TLR2- and TLR4-dependent as demonstrated with TLR-transfected fibroblastoid cells, as well as macrophages and dendritic cells from functional TLR-deficient mice; 3) blood cells of onchocerciasis patients exposed to rWSP also generated down-regulating mediators IL-10 and PGE2 after 6 days of culture; 4) furthermore, rWSP-reactive IgG1 Abs were present in sera of O. volvulus-infected people but not in those of uninfected Europeans. The lack of rWSP-reactive IgE and IgG4 in serum indicated a bias toward a Th1-type adaptive immune response. Abs against rWSP stained endobacteria in living and degenerating adult O. volvulus filariae, tissue microfilariae and host tissue macrophages that apparently had engulfed microfilariae. Thus, filarial helminths, through products of their endobacteria such as WSP, acquire characteristics of a typical microbial pathogen inducing immune responses via TLR2 and TLR4.

cDNA cloning of turtle prion protein

Cloning of the cDNA coding for the 270‐residue turtle prion protein is reported. It represents the most remote example thus far described. The entire coding region is comprised in a single exon, while a large intron interrupts the 5′ UTR. The common structural features of the known prion proteins are all conserved in turtle PrP, whose identity degree to mammalian and avian proteins is about 40 and 58%, respectively. The most intriguing feature, unique to the turtle prion, is the presence of an EF‐hand Ca2+ binding motif in the C‐terminal half of the protein.

Antigenic role of the endosymbionts of filarial nematodes: IgG response against the Wolbachia surface protein in cats infected with Dirofilaria immitis

Filarial nematodes harbour intracellular endosymbiotic bacteria, which have been assigned to the genus Wolbachia. These bacteria appear to play an important role in the pathogenesis of filarial diseases through their lipopolysaccharides. In view of the presence of Wolbachia endosymbionts in the body of filarial nematodes, one might also expect that proteins from these bacteria play an antigenic role in humans and animals affected by filariases. To test this hypothesis, we produced in recombinant form the surface protein WSP and a portion of the cell–cycle protein FTSZ from the Wolbachia of Dirofilaria immitis. Western immunoblot assays were then performed using cat sera to test the immunogenicity of these proteins. Sera were collected from owners cats, which were either sero–negative or sero–positive for D.immitis and from cats before and after experimental infection with D.immitis. FTSZ was recognized in Western blots by sera from both positive and negative cats and from both uninfected and experimentally infected cats.WSP was recognized only by sera from positive cats and from cats experimentally infected with D.immitis; this protein was not recognized by sera from negative cats and from cats before experimental infection with D.immitis. The results of Western blot assays on WSP thus support the hypothesis that infection with filarial nematodes induces the production of antibodies against Wolbachia proteins.

Animal board invited review: advances in proteomics for animal and food sciences

Animal production and health (APH) is an important sector in the world economy, representing a large proportion of the budget of all member states in the European Union and in other continents. APH is a highly competitive sector with a strong emphasis on innovation and, albeit with country to country variations, on scientific research. Proteomics (the study of all proteins present in a given tissue or fluid – i.e. the proteome) has an enormous potential when applied to APH. Nevertheless, for a variety of reasons and in contrast to disciplines such as plant sciences or human biomedicine, such potential is only now being tapped. To counter such limited usage, 6 years ago we created a consortium dedicated to the applications of Proteomics to APH, specifically in the form of a Cooperation in Science and Technology (COST) Action, termed FA1002 – Proteomics in Farm Animals: www.cost-faproteomics.org. In 4 years, the consortium quickly enlarged to a total of 31 countries in Europe, as well as Israel, Argentina, Australia and New Zealand. This article has a triple purpose. First, we aim to provide clear examples on the applications and benefits of the use of proteomics in all aspects related to APH. Second, we provide insights and possibilities on the new trends and objectives for APH proteomics applications and technologies for the years to come. Finally, we provide an overview and balance of the major activities and accomplishments of the COST Action on Farm Animal Proteomics. These include activities such as the organization of seminars, workshops and major scientific conferences, organization of summer schools, financing Short-Term Scientific Missions (STSMs) and the generation of scientific literature. Overall, the Action has attained all of the proposed objectives and has made considerable difference by putting proteomics on the global map for animal and veterinary researchers in general and by contributing significantly to reduce the East–West and North–South gaps existing in the European farm animal research. Future activities of significance in the field of scientific research, involving members of the action, as well as others, will likely be established in the future.

Swarming Differentiation and Swimming Motility in Bacillus subtilis Are Controlled by swrA, a Newly Identified Dicistronic Operon

The number and disposition of flagella harbored by eubacteria are regulated by a specific trait successfully maintained over generations. The genes governing the number of flagella in Bacillus subtilis have never been identified, although the ifm locus has long been recognized to influence the motility phenotype of this microorganism. The characterization of a spontaneous ifm mutant of B. subtilis, displaying diverse degrees of cell flagellation in both liquid and solid media, raised the question of how the ifm locus governs the number and assembly of functional flagella. The major finding of this investigation is the characterization of a newly identified dicistronic operon, named swrA, that controls both swimming motility and swarming differentiation in B. subtilis. Functional analysis of the swrA operon allowed swrAA (previously named swrA [D. B. Kearns, F. Chu, R. Rudner, and R. Losick, Mol. Microbiol. 52:357-369, 2004]) to be the first gene identified in B. subtilis that controls the number of flagella in liquid environments and the assembly of flagella in response to cell contact with solid surfaces. Evidence is given that the second gene of the operon, swrAB, is essential for enabling the surface-adhering cells to undergo swarming differentiation. Preliminary data point to a molecular interaction between the two gene products.

The primary structure of UK 114 tumor antigen

UK114 is a tumor antigen expressed by various malignant neoplasms. The complete amino acid sequence of UK114 purified from goat liver has been determined by automated Edman degradation of CNBr and endoproteinase Lys‐C peptides. The protein contains 137 amino acid residues, which corresponds to a molecular mass of 14 229 Da. MALDITOF analysis resulted in a molecular weight of 14 290, suggesting that the N‐terminal Met residue is acetylated. Sequence comparison shows that UK114 from goat liver (1) has 77% identity with a previously described 23 kDa protein from rat liver (Levy‐Favatier et al. (1993) Eur. J. Biochem. 212, 665–673), (2) shares a very high degree of similarity with a family of prokaryotic and eukaryotic hypothetic proteins whose function have not yet been characterized, and (3) exhibits a significant similarity to a group of tumor‐associated antigens which belongs to a superfamily of heat shock proteins, acting as possible targets for the hosts antitumor immunity.

Purification, inhibitory properties, amino acid sequence and identification of the reactive site of a new serine proteinase inhibitor from oil‐rape (itBrassica napus) seed

A new serine proteinase inhibitor, rapeseed trypsin inhibitor (RTI), has been isolated from rapeseed (Brassica napus var. oleifera) seed. The protein inhibits the catalytic activity of bovine β‐trypsin and bovine α‐chymotrypsin with apparent dissociation constants of 3.0 × 10−10 M and 4.1 × 10−7 M, at pH 8.0 and 21°C, respectively. The stoichiometry of both proteinase‐inhibitor complexes is 1:1. The amino acid sequence of RTI consists of 60 amino acid residues, corresponding to an M r, of about 6.7 kDa. The p1‐pi, reactive site bond has been tentatively identified at position Arg20‐Ile21. RTI shows no similarity to other serine proteinase inhibitors except the low molecular weight mustard trypsin inhibitor (MTI‐2). RTI and MTI‐2 could be members of a new class of plant serine proteinase inhibitors.

The primary structure of D-amino acid oxidase from Rhodotorula gracilis

SummaryThe amino acid sequence of D-amino acid oxidase from Rhodotorula gracilis was determined by automated Edman degradation of peptides generated by enzymatic and chemical cleavage. The enzyme monomer contains 368 amino acid residues and its sequence is homologous to that of other known D-amino acid oxidases. Six highly conserved regions appear to have a specific role in binding of coenzyme FAD, in active site topology and in peroxisomal targeting. Moreover, Rhodotorula gracilis D-amino acid oxidase contains a region with a cluster of basic amino acids, probably exposed to solvent, which is absent in other D-amino acid oxidases.