Evidence of a Lytic Pathway in an Invertebrate Complement System: Identification of a Terminal Complement Complex Gene in a Colonial Tunicate and Its Evolutionary Implications.

The complement system is a pivotal component of innate immunity, extensively studied in vertebrates but also present in invertebrates. This study explores the existence of a terminal complement pathway in the tunicate , aiming to understand the evolutionary integration of innate and adaptive immunity. Through transcriptome analysis, we identified a novel transcript, BsITCCP, encoding a protein with both MACPF and LDLa domains-a structure resembling that of vertebrate C9 but with a simpler organization.

Clodronate Liposome-Mediated Phagocytic Hemocyte Depletion Affects the Regeneration of the Cephalic Tentacle of the Invasive Snail, .

After amputation, granular hemocytes infiltrate the blastema of regenerating cephalic tentacles of the freshwater snail . Here, the circulating phagocytic hemocytes were chemically depleted by injecting the snails with clodronate liposomes, and the effects on the cephalic tentacle regeneration onset and on -Hemocyanin, -transglutaminase (-TG) and -Allograft Inflammatory Factor-1 (-AIF-1) gene expressions were investigated. Flow cytometry analysis demonstrated that clodronate liposomes targeted large circulating hemocytes, resulting in a transient decrease in their number.

The Eco-Immunological Relevance of the Anti-Oxidant Response in Invasive Molluscs.

Reactive oxygen species (ROS) are volatile and short-lived molecules playing important roles in several physiological functions, including immunity and physiological adaptation to unsuitable environmental conditions. In an eco-immunological view, the energetic costs associated with an advantageous metabolic apparatus able to cope with wide changes in environmental parameters, e.g.

Botryllin, a Novel Antimicrobial Peptide from the Colonial Ascidian .

By mining the transcriptome of the colonial ascidian , we identified a transcript for a novel styelin-like antimicrobial peptide, which we named botryllin. The gene is constitutively transcribed by circulating cytotoxic morula cells (MCs) as a pre-propeptide that is then cleaved to mature peptide. The synthetic peptide, obtained from in silico translation of the transcript, shows robust killing activity of bacterial and unicellular yeast cells, causing breakages of both the plasma membrane and the cell wall.

The echinoid complement system inferred from genome sequence searches.

The vertebrate complement cascade is an essential host protection system that functions at the intersection of adaptive and innate immunity. However, it was originally assumed that complement was present only in vertebrates because it was activated by antibodies and functioned with adaptive immunity. Subsequently, the identification of the key component, SpC3, in sea urchins plus a wide range of other invertebrates significantly expanded the concepts of how complement functions.

Stress granules in Ciona robusta: First evidences of TIA-1-related nucleolysin and tristetraprolin gene expression under metal exposure.

Stress granules are non-membranous cytoplasmic foci, composed of non-translating messenger ribonucleoproteins, translational initiation factors and other additional proteins. They represent a primary mechanism to rapidly modulate gene expression when cells are subjected to adverse environmental conditions. Very few works have been devoted to study the presence of the molecular components of stress granules in invertebrates.

BsTLR1: A new member of the TLR family of recognition proteins from the colonial ascidian Botryllus schlosseri.

Toll-like receptors (TLRs) represent a well-known family of conserved pattern recognition receptors the importance of which, in non-self recognition, was demonstrated in both vertebrates and invertebrates. Tunicates represent the vertebrate sister group and, as invertebrates, they rely only on innate immunity for their defence. As regards TLRs, two transcripts have been described and characterised in the solitary species Ciona intestinalis, referred to as CiTLR1 and CiTLR2.

Insights into the Complement System of Tunicates: C3a/C5aR of the Colonial Ascidian .

As an evolutionary ancient component of the metazoan immune defense toolkit, the complement system can modulate cells and humoral responses of both innate and (in jawed vertebrates) adaptive immunity. All the three known complement-activation pathways converge on the cleavage of C3 to C3a and C3b. The anaphylatoxin C3a behaves as a chemokine in inflammatory responses, whereas C3b exerts an opsonic role and, ultimately, can activate the lytic pathway.

Complement system and phagocytosis in a colonial protochordate.

In the present work, we investigated, in the colonial ascidian Botryllus schlosseri, the role of complement C3 (BsC3) in phagocytosis. We studied the modulation of BsC3 transcription in the course of the colonial blastogenetic cycle, with particular reference to the takeover, when apoptotic cells in the tissues of old zooids are cleared by circulating phagocytes. In situ hybridisation with BsC3 riboprobes labelled only morula cells, the most abundant haemocytes.

Functional amyloidogenesis in immunocytes from the colonial ascidian Botryllus schlosseri: Evolutionary perspective.

Cytotoxic morula cells (MCs) and phagocytes are the circulating immunocytes of the colonial ascidian Botryllus schlosseri: Both these cells can synthesise amyloid fibrils, supporting the idea that physiological amyloidogenesis is involved in inflammation and modulation of immune responses. Intriguingly, amyloid of B. schlosseri immunocytes is made of two different proteins.

Immunity in Protochordates: The Tunicate Perspective.

Tunicates are the closest relatives of vertebrates, and their peculiar phylogenetic position explains the increasing interest toward tunicate immunobiology. They are filter-feeding organisms, and this greatly influences their defense strategies. The majority of the studies on tunicate immunity were carried out in ascidians.

Protection from Oxidative Stress in Immunocytes of the Colonial Ascidian Botryllus schlosseri: Transcript Characterization and Expression Studies.

Botryllus schlosseri is a cosmopolitan colonial ascidian that undergoes cyclical generation changes, or take-overs, during which adult zooids are resorbed and replaced by their buds. At take-over, adult tissues undergo diffuse apoptosis and effete cells are massively ingested by circulating phagocytes, with a consequent increase in oxygen consumption and in production of reactive oxygen species (ROS). The latter are responsible for the death of phagocytes involved in the clearance of apoptotic cells and corpses by phagocytosis-induced apoptosis.

Morula cells as key hemocytes of the lectin pathway of complement activation in the colonial tunicate Botryllus schlosseri.

The complement system is deeply rooted in the evolution of humoral mechanism of innate immunity. In addition to the alternative pathway of complement activation, lectins and associated serine proteases exert important roles in the recognition of non-self and activation of the effectors. In the colonial tunicate Botryllus schlosseri, we identified, characterized and studied the expression of three orthologues of genes involved in the lectin pathway of complement activation of vertebrates, i.

Data on four apoptosis-related genes in the colonial tunicate Botryllus schlosseri.

The data described are related to the article entitled "Recurrent phagocytosis-induced apoptosis in the cyclical generation change of the compound ascidian Botryllus schlosseri" (Franchi et al., 2016) [1]. Four apoptosis-related genes, showing high similarity with mammalian Bax (a member of the Bcl-2 protein family), AIF1 (apoptosis-inducing factor-1), PARP1 (poly ADP ribose polymerase-1) and IAP7 (inhibitor of apoptosis-7) were identified from the analysis of the trascriptome of B.

Histamine Stimulates Ciliary Beat Frequency via the H2 Receptor in the Protochordate Botryllus schlosseri.

Histamine is a biogenic molecule that plays a role in many physiological pathways via binding to a specific receptor. Histaminergic receptors belong to the large family of seven-transmembrane α-helix domain receptors classified in mammals into four distinct classes: H1, H2, H3, and H4. Despite being widely studied in vertebrates, few data are available on the invertebrate receptors, with only predicted H1 and H2 sequences for nonchordate deuterostomes.

Recurrent phagocytosis-induced apoptosis in the cyclical generation change of the compound ascidian Botryllus schlosseri.

Colonies of the marine, filter-feeding ascidian Botryllus schlosseri undergo cyclical generation changes or takeovers. These events are characterised by the progressive resorption of adult zooids and their replacement by their buds that grow to adult size, open their siphons and start filtering. During the take-over, tissues of adult zooids undergo extensive apoptosis; circulating, spreading phagocytes enter the effete tissues, ingest dying cells acquiring a giant size and a round morphology.

Transcriptome dynamics in the asexual cycle of the chordate Botryllus schlosseri.

Background: We performed an analysis of the transcriptome during the blastogenesis of the chordate Botryllus schlosseri, focusing in particular on genes involved in cell death by apoptosis. The tunicate B. schlosseri is an ascidian forming colonies characterized by the coexistence of three blastogenetic generations: filter-feeding adults, buds on adults, and budlets on buds.

SATRAP: SOLiD Assembler TRAnslation Program.

SOLiD DNA sequences are typically analyzed using a reference genome, while they are not recommended for de novo assembly of genomes or transcriptomes. This is mainly due to the difficulty in translating the SOLiD color-space data into normal base-space sequences. In fact, the nature of color-space is such that any misinterpreted color leads to a chain of further translation errors, producing totally wrong results.

Sexual and asexual reproduction in the colonial ascidian Botryllus schlosseri.

The colonial tunicate Botryllus schlosseri is a widespread filter-feeding ascidian that lives in shallow waters and is easily reared in aquaria. Its peculiar blastogenetic cycle, characterized by the presence of three blastogenetic generations (filtering adults, buds, and budlets) and by recurrent generation changes, has resulted in over 60 years of studies aimed at understanding how sexual and asexual reproduction are coordinated and regulated in the colony. The possibility of using different methodological approaches, from classical genetics to cell transplantation, contributed to the development of this species as a valuable model organism for the study of a variety of biological processes.

Preliminary characterization of complement in a colonial tunicate: C3, Bf and inhibition of C3 opsonic activity by compstatin.

The complement system is a fundamental effector mechanism of the innate immunity in both vertebrates and invertebrates. The comprehension of its roots in the evolution is a useful step to understand how the main complement-related proteins had changed in order to adapt to new environmental conditions and life-cycles or, in the case of vertebrates, to interact with the adaptive immunity. Data on organisms evolutionary close to vertebrates, such as tunicates, are of primary importance for a better understanding of the changes in immune responses associated with the invertebrate-vertebrate transition.

Characterization and transcription studies of a phytochelatin synthase gene from the solitary tunicate Ciona intestinalis exposed to cadmium.

The major thiol-containing molecules involved in controlling the level of intracellular ROS in eukaryotes, acting as a nonenzymatic detoxification system, are metallothioneins (MTs), glutathione (GSH) and phytochelatins (PCs). Both MTs and GSH are well-known in the animal kingdom. PC was considered a prerogative of the plant kingdom but, in 2001, a phytochelatin synthase (PCS) gene was described in the nematode Caenorhabditis elegans; additional genes encoding this enzyme were later described in the earthworm Eisenia fetida and in the parasitic nematode Schistosoma mansoni but scanty data are available, up to now, for Deuterostomes.

In vitro effects of the nonsteroidal anti-inflammatory drug, ibuprofen, on the immune parameters of the colonial ascidian Botryllus schlosseri.

In this study, in vitro effects of ibuprofen (IBU) on the immune parameters of the colonial ascidian Botryllus schlosseri were evaluated. Haemocytes were exposed for 1h to 0 (control), 100 and 1000 μg IBU/L and the effects on haemocyte viability and morphology (shape factor), lysosomal membrane stability (Neutral Red Retention Assay), phagocytic activity, apoptosis (TUNEL reaction), hydrolytic (acid phosphatase) and oxidative (phenoloxidase and peroxidase) enzyme activities were evaluated. The exposure of haemocytes to IBU did not affect significantly their viability, but increased the percentage of cells with round shape.

Characterization and metal-induced gene transcription of two new copper zinc superoxide dismutases in the solitary ascidian Ciona intestinalis.

Antioxidant enzymes are known to protect living organisms against the oxidative stress risk, also induced by metals. In the present study, we describe the purification and molecular characterization of two Cu,Zn superoxide dismutases (SODs), referred to as Ci-SODa and Ci-SODb, from Ciona intestinalis, a basal chordate widely distributed in temperate shallow seawater. The putative amino acid sequences were compared with Cu,Zn SODs from other metazoans and phylogenetic analyses indicate that the two putative Ci-SODs are more related to invertebrate SODs than vertebrate ones.

Influence of cadmium on the morphology and functionality of haemocytes in the compound ascidian Botryllus schlosseri.

In order to get insights into the effects of cadmium (Cd) on cell morphology and functions, we exposed haemocytes of the colonial ascidian Botryllus schlosseri to sub-lethal concentrations of CdCl(2). Results indicate that Cd hampers haemocyte spreading and phagocytosis in a dose-dependent way, through the alteration of the actin cytoskeleton. In addition, the metal decreases the stability of the internal membranes, as revealed by the Neutral Red assay.

Insight on signal transduction pathways involved in phagocytosis in the colonial ascidian Botryllus schlosseri.

Tunicates are chordate invertebrates, closely related to vertebrates, which represent valuable organisms for the study of a variety of biological processes from an evolutionary point of view. As invertebrates, they rely on innate immunity to cope with foreign, potentially pathogenic material. Among tunicates, the compound ascidian Botryllus schlosseri is emerging as a reliable model organism for the study of innate immune responses.