Research on innate immunity of the penaeid shrimps and the oyster Crassostrea gigas is motivated greatly by economical necessities. Indeed, the aquaculture of these organisms is now limited by the development of infectious diseases. Studying anti-microbial peptides/proteins (AMPs), which are effector molecules of the host defense, is particularly attractive not only for progressing basic knowledge on immunity but also because they offer various possible applications for disease management in aquaculture. AMPs are explored with a global approach,considering their structure, properties, function, gene expression, and tissue distribution during the response to infections. In shrimp, investigations of the penaeidins, which are constitutively expressed peptides, have highlighted the importance of hemocytes and hematopoiesis as major elements of the immune response, providing both local and systemic reactions. The activation of hematopoiesis must be regarded as a regulatory way for the expression and distribution of constitutively expressed immune effectors. As complementary approaches, genomics and gene profiling are promising to deepen our understanding of the anti-microbial defense of the oyster and the shrimp. However, real progress will depend also on the characterization of hemocyte lineages and hematopoiesis of these marine invertebrates as well as on the ontogenesis of their immune systems.

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Keratinocyte differentiation program leading to an organized epidermis plays a key role in maintaining the first line of defense of the skin. Epidermal integrity is regulated by a tight communication between keratinocytes and leucocytes, particularly under cytokine control. Imbalance of the cytokine network leads to inflammatory diseases such as psoriasis. Our attempt to model skin inflammation showed that the combination of IL-17A, IL-22, IL-1α, OSM and TNFα (Mix M5) synergistically increases chemokine and antimicrobial-peptide expression, recapitulating some features of psoriasis. Other characteristics of psoriasis are acanthosis and down-regulation of keratinocyte differentiation markers. Our aim was to characterize the specific roles of these cytokines on keratinocyte differentiation, and to compare with psoriatic lesion features. All cytokines decrease keratinocyte differentiation markers, but IL-22 and OSM were the most powerful, and the M5 strongly synergized the effects. In addition, IL-22 and OSM induced epidermal hyperplasia in vitro and M5 induced epidermal thickening and decreased differentiation marker expression in a mouse model, as observed in human psoriatic skin lesions. This study highlights the precise role of cytokines in the skin inflammatory response. IL-22 and OSM more specifically drive epidermal hyperplasia and differentiation loss while IL-1α, IL-17A and TNFα were more involved in the activation of innate immunity.

Antimicrobial peptides play a major role in innate immunity. The penaeidins, initially characterized from the shrimp Litopenaeus vannamei, are a family of antimicrobial peptides that appear to be expressed in all penaeid shrimps. As of recent, a large number of penaeid nucleotide sequences have been identified from a variety of penaeid shrimp species and these sequences currently reside in several databases under unique identifiers with no nomenclatural continuity. To facilitate research in this field and avoid potential confusion due to a diverse number of nomenclatural designations, we have made a systematic effort to collect, analyse, and classify all the penaeidin sequences available in every database. We have identified a common penaeidin signature and subsequently established a classification based on amino acid sequences. In order to clarify the naming process, we have introduced a 'penaeidin nomenclature' that can be applied to all extant and future penaeidins. A specialized database, PenBase, which is freely available at , has been developed for the penaeidin family of antimicrobial peptides, to provide comprehensive information about their properties, diversity and nomenclature.

In invertebrates, defensins were found in arthropods and in the mussels. Here, we report for the first time the identification and characterization of a defensin (Cg-Def) from an oyster. Cg-def mRNA was isolated from Crassostrea gigas mantle using an expressed sequence tag approach. To gain insight into potential roles of Cg-Def in oyster immunity, we produced the recombinant peptide in Escherichia coli, characterized its antimicrobial activities, determined its solution structure by NMR spectroscopy, and quantified its gene expression in vivo following bacterial challenge of oysters. Recombinant Cg-Def was active in vitro against Grampositive bacteria but showed no or limited activities against Gramnegative bacteria and fungi. The activity of Cg-Def was retained in vitro at a salt concentration similar to that of seawater. The Cg-Def structure shares the so-called cystine-stabilized ␣-␤ motif (CS-␣␤) with arthropod defensins but is characterized by the presence of an additional disulfide bond, as previously observed in the mussel defensin (MGD-1). Nevertheless, despite a similar global fold, the Cg-Def and MGD-1 structures mainly differ by the size of their loops and by the presence of two aspartic residues in Cg-Def. Distribution of Cg-def mRNA in various oyster tissues revealed that Cgdef is mainly expressed in mantle edge where it was detected by mass spectrometry analyses. Furthermore, we observed that the Cg-def messenger concentration was unchanged after bacterial challenge. Our results suggest that Cg-def gene is continuously expressed in the mantle and would play a key role in oyster by providing a first line of defense against pathogen colonization. Antimicrobial peptides (AMPs)5 are important components of the innate immune system that have been conserved during evolution (1). They constitute a first line of host defense against pathogens in plants and animals (2, 3). We estimate that more than 1000 antimicrobial peptides have been described at the level of their primary structure (2). They are gathered in the Antimicrobial Sequence Database (www.bbcm.units.it/ϳtossi/amsdb.html). AMPs can be classified into three major groups: (i) linear peptides that form amphipathic ␣-helices, (ii) cyclic peptides containing cysteine-residue engaged in disulfide bonds, and (iii) peptides with an overrepresentation in certain amino acids (proline, arginine, glycine, or histidine). Despite their great diversity in terms of size, primary structure, amino acid composition, and mode of action, most AMPs are characterized by the preponderance in cationic and hydrophobic amino acids (2). In most of the cases, this amphipathic character is considered as crucial for the interaction of the effective peptide with the membrane of sensitive microorganisms. This first interaction seems to be essential whatever the exact mode of action: (i) through disruption of their negatively charged cytoplasmic membranes or (ii) through killing following translocation into the bacteria without membrane lyses and binding to a specific target pr...

Cutaneous homeostasis and defenses are maintained by permanent cross-talk among particular epidermal keratinocytes and immune cells residing or recruited in the skin, through the production of cytokines. If required, a coordinated inflammatory response is triggered, relayed by specific cytokines. Due to numerous reasons, troubles in the resolution of this phenomenon could generate a cytokine-mediated vicious circle, promoting skin chronic inflammation, the most common being atopic dermatitis and psoriasis. In this paper, we discuss the biological effects of cytokine on keratinocytes, more particularly on specific or shared cytokines involved in atopic dermatitis or psoriasis. We report and discuss monolayer or 3D in vitro models of keratinocytes stimulated by specific sets of cytokines to mimic atopic dermatitis or psoriasis. IL-22, TNFa, IL-4, and IL-13 combination is able to mimic an “atopic dermatitis like” state. In psoriasis lesions, over expression of IL-17 is observed whereas IL-4 and IL-13 were not detected; the replacement of IL-4 and IL-13 by IL-17 from this mix is able to mimic in vitro a “psoriasis like” status on keratinocytes. We conclude that specific cytokine environment deregulation plays a central role on skin morphology and innate immunity, moving towards specific pathologies and opening the way to new therapeutic strategies.

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Penaeidins are a family of antimicrobial peptides of 47-63 residues isolated from several species of shrimp. These peptides display a proline-rich domain (N-terminal part) and a cysteine-rich domain (C-terminal part) stabilized by three conserved disulfide bonds whose arrangement has not yet been characterized. The recombinant penaeidin-3a of Litopenaeus vannamei (63 residues) and its [T8A]-Pen-3a analogue were produced in Saccharomyces cerevisiae and showed similar antimicrobial activity. The solution structure of the [T8A]-Pen3a analogue was determined by using two-dimensional 1 H NMR and simulated annealing calculations. The proline-rich domain, spanning residues 1-28 was found to be unconstrained. In contrast, the cysteine-rich domain, spanning residues 29 -58, displays a well defined structure, which consists of an amphipathic helix (41-50) linked to the upstream and the downstream coils by two disulfide bonds (Cys 32 -Cys 47 and Cys 48 -Cys 55 ). These two coils are in turn linked together by the third disulfide bond (Cys 36 -Cys 54 ). Such a disulfide bond packing, which is in agreement with the analysis of trypsin digests by ESI-MS, contributes to the highly hydrophobic core. Side chains of Arg 45 and Arg 50 , which belong to the helix, and side chains of Arg 37 and Arg 53 , which belong to the upstream and the downstream coils, are located in two opposite parts of this globular and compact structure. The environment of these positively charged residues, either by hydrophobic clusters at the surface of the cysteine-rich domain or by sequential hydrophobic residues in the unconstrained proline-rich domain, gives rise to the amphipathic character required for antimicrobial peptides. We hypothesize that the antimicrobial activity of penaeidins can be explained by a cooperative effect between the proline-rich and cysteine-rich features simultaneously present in their sequences.Antimicrobial peptides are major elements of host-defense systems represented in all species from plants to vertebrate and invertebrate animals. Among these antimicrobial molecules, cysteine-rich peptides are the most widespread. They are structurally classified into (i) peptides with a ␤-sheet structure such as the mammalian defensins (1), (ii) peptides with a ␤-hairpin-like fold such as tachyplesins from horseshoe crabs (2), thanatin (3), porcine protegrins (4, 5), androctonin (6), or gomesin (7), and (iii) peptides adopting the cystine-stabilized ␣␤-motif, including invertebrate and plant defensins (8Ϫ10).Only recently, such effectors of innate immunity were isolated from crustaceans, whereas numerous peptides have been characterized from other arthropods, both insects and chelicerates. Three peptides, named penaeidins (Pen), 1 were initially purified in their active form (5.48Ϫ6.62 kDa) from the hemocytes of the shrimp Litopenaeus (Penaeus) vannamei, and they were fully characterized at the amino acid level (Pen-1, Pen-2, Pen3a) and by cDNA cloning from a hemocyte library (pen-2, -3a, -3b, and-3c) (11). Penaeidins are an original p...