Variable domains in hagfish: NICIR is a polymorphic multigene family expressed preferentially in leukocytes and is related to lamprey TCR-like

The jawless vertebrates, represented by hagfish and lampreys, are the most advanced animals that apparently lack T cell and B cell receptors. As such, they offer unique opportunities for understanding the evolution of antigen receptors and variable (V)-type immunoglobulin (Ig)-like domains. In the present study, we describe four hagfish Ig superfamily (IgSF) members carrying V-type domains. None of them appeared to have direct counterparts in jawed vertebrates, indicating that many IgSF molecules have either evolved independently in jawed and jawless vertebrates or diverged to the extent that clear homology is no longer recognizable. One of the members encoded a molecule closely related to the previously described membrane protein designated novel ITAM (immunoreceptor tyrosine-based activation motif)-containing IgSF receptors (NICIR). We show here that NICIR is a polymorphic multigene family with at least three members and is expressed predominantly in peripheral blood leukocytes. Phylogenetic analysis indicates that among known proteins, NICIR is most closely related to the lamprey molecule recently proposed to be a potential ancestor of T cell receptors.Sequence data reported in this paper were submitted to the DDBJ/EMBL/GenBank databases under accession nos. AB234206-AB234210, AB242215-AB224219, and AB242221-AB242223.     

Structure of Signal-regulatory Protein ��: A LINK TO ANTIGEN RECEPTOR EVOLUTION*

Signal-regulatory protein α (SIRPα) is a myeloid membrane receptor that interacts with the membrane protein CD47, a marker of self. We have solved the structure of the complete extracellular portion of SIRPα, comprising three immunoglobulin superfamily domains, by x-ray crystallography to 2.5 Å resolution. These data, together with previous data on the N-terminal domain and its ligand CD47 (possessing a single immunoglobulin superfamily domain), show that the CD47-SIRPα interaction will span a distance of around 14 nm between interacting cells, comparable with that of an immunological synapse. The N-terminal (V-set) domain mediates binding to CD47, and the two others are found to be constant (C1-set) domains. C1-set domains are restricted to proteins involved in vertebrate antigen recognition: T cell antigen receptors, immunoglobulins, major histocompatibility complex antigens, tapasin, and β2-microglobulin. The domains of SIRPα (domains 2 and 3) are structurally more similar to C1-set domains than any cell surface protein not involved in antigen recognition. This strengthens the suggestion from sequence analysis that SIRP is evolutionarily closely related to antigen recognition proteins.Signal-regulatory protein α (SIRPα)⁴ is a membrane receptor present on myeloid cells and neurons that interacts with the widely distributed cell surface protein CD47 (reviewed in Refs. 1 and 2). Absence of CD47 leads to uptake of cells via macrophages, indicating that CD47 acts as a marker of self (3). SIRPα gives inhibitory signals through immunoreceptor tyrosine-based inhibition motifs in the cytoplasmic region that interact with phosphatases SHP-1 and SHP-2 (4). Binding of the N-terminal immunoglobulin superfamily (IgSF) V-set domain of SIRPα (SIRPα d1) to the single IgSF domain of CD47 is mediated by the loops of the SIRPα IgSF domain, analogous to the interactions mediated by antigen receptors, albeit involving only a single domain (5, 6). This type of binding distinguishes the CD47-SIRPα interaction from that of many interactions at the cell surface involving IgSF domains such as CD2-CD58, where the face of the IgSF domain is involved (7). SIRPα domains 2 and 3 (d2 and d3) show amino acid sequence similarity to IgSF C1-set domains (8). Since IgSF C1-set domains have only been confirmed in vertebrate antigen receptors and associated proteins (Ig light and heavy chains, T cell receptor chains, MHC class I and II and related proteins, β2-microglobulin, and very recently tapasin (9)) of the vertebrate adaptive immune system, it was suggested that SIRPα might have evolved from a precursor of the antigen receptors (8).We describe here the crystal structure of the full three-domain extracellular region of SIRPα, revealing that the topology of the CD47-SIRPα interaction is compatible with productive engagement occurring when cells come together in synapse-like contacts. We show that the two membrane-proximal IgSF domains are particularly close in structure to C1-set IgSF domains. This, together with the presence of an IgSF V-set domain mediating ligand recognition, suggests that SIRPα is related to a key precursor in the evolution of vertebrate antigen receptors.     

A novel soluble immune-type receptor (SITR) in teleost fish: carp SITR is involved in the nitric oxide-mediated response to a protozoan parasite

Background

The innate immune system relies upon a wide range of germ-line encoded receptors including a large number of immunoglobulin superfamily (IgSF) receptors. Different Ig-like immune receptor families have been reported in mammals, birds, amphibians and fish. Most innate immune receptors of the IgSF are type I transmembrane proteins containing one or more extracellular Ig-like domains and their regulation of effector functions is mediated intracellularly by distinct stimulatory or inhibitory pathways.

Methodology/Principal Findings

Carp SITR was found in a substracted cDNA repertoire from carp macrophages, enriched for genes up-regulated in response to the protozoan parasite Trypanoplasma borreli. Carp SITR is a type I protein with two extracellular Ig domains in a unique organisation of a N-proximal V/C2 (or I-) type and a C-proximal V-type Ig domain, devoid of a transmembrane domain or any intracytoplasmic signalling motif. The carp SITR C-proximal V-type Ig domain, in particular, has a close sequence similarity and conserved structural characteristics to the mammalian CD300 molecules. By generating an anti-SITR antibody we could show that SITR protein expression was restricted to cells of the myeloid lineage. Carp SITR is abundantly expressed in macrophages and is secreted upon in vitro stimulation with the protozoan parasite T. borreli. Secretion of SITR protein during in vivo T. borreli infection suggests a role for this IgSF receptor in the host response to this protozoan parasite. Overexpression of carp SITR in mouse macrophages and knock-down of SITR protein expression in carp macrophages, using morpholino antisense technology, provided evidence for the involvement of carp SITR in the parasite-induced NO production.

Conclusion/Significance

We report the structural and functional characterization of a novel soluble immune-type receptor (SITR) in a teleost fish and propose a role for carp SITR in the NO-mediated response to a protozoan parasite.     

Characterization of the laminin binding domains of the Lutheran blood group glycoprotein

Lutheran (Lu) blood group antigens and the basal cell adhesion molecule antigen reside on two glycoproteins that belong to the Ig superfamily (IgSF) and carry five Ig-like extracellular domains. These glycoproteins act as widely expressed adhesion molecules and represent the unique receptors for laminin-10/11 in erythroid cells. Here, we report the mapping of IgSF domains responsible for binding to laminin. In plasmonic resonance surface experiments, only recombinant Lu proteins containing the N-terminal IgSF domains 1-3 were able to bind laminin-10/11 and to inhibit binding of laminin to Lu-expressing K562 cells. Mutant recombinant proteins containing only IgSF domain 1, domains 1 + 2, domains 1 + 3, domains 2 + 3, domain 3, domain 4, domain 5, and domains 4 + 5 failed to bind laminin as well as a construct containing all of the extracellular domains except domain 3. Altogether, these results indicate that IgSF domains 1-3 are involved in laminin binding and that a specific spatial arrangement of these three first domains is most probably necessary for interaction. Neither the RGD nor the N-glycosylation motifs present in IgSF domain 3 were involved in laminin binding.     

The immunoglobulin superfamily: An insight on its tissular, species, and functional diversity

The immunoglobulin superfamily (IgSF) is a heterogenic group of proteins built on a common fold, called the Ig fold, which is a sandwich of two β sheets. Although members of the IgSF share a similar Ig fold, they differ in their tissue distribution, amino acid composition, and biological role. In this paper we report an up-to-date compilation of the IgSF where all known members of the IgSF are classified on the basis of their common functional role (immune system, antibiotic proteins, enzymes, cytokine receptors, etc.) and their distribution in tissue (neural system, extracellular matrix, tumor marker, muscular proteins, etc.), or in species (vertebrates, invertebrates, bacteria, viruses, fungi, and plants). The members of the family can contain one or many Ig domains, comprising two basic types: the constant domain (C), with seven strands, and the variable domain (V), with eight, nine, or ten strands. The different overviews of the IgSF led to the definition of new domain subtypes, mainly concerning the C type, based on the distribution of strands within the two sheets. The wide occurrence of the Ig fold and the much less conserved sequences could have developed from a common ancestral gene and/or from a convergent evolutionary process. Cell adhesion and pattern recognition seem to be the common feature running through the entire family. Received: 4 June 1997 / Accepted: 15 September 1997     

Structural Evidence for the Evolution of Pyrogenic Toxin Superantigens

Cells from metazoan organisms are eliminated in a variety of physiological and pathophysiological processes by apoptosis. In this report, we describe the cloning and characterization of molecules from the marine sponges Geodia cydonium and Suberites domuncula, whose domains show a high similarity to those that are found in molecules of the vertebrate Bcl-2 superfamily and of the death receptors. The Bcl-2 proteins contain up to four Bcl-2 homology regions (BH). Two Bcl-2-related molecules have been identified from sponges that are provided with two of those regions, BH1 and BH2, and are termed Bcl-2 homology proteins (BHP). The G. cydonium molecule, BHP1_GC, has a putative size of 28,164, while the related sequence from S. domuncula, BHP1_SD, has a M _r of 24,187. Phylogenetic analyses of the entire two sponge BHPs revealed a high similarity to members of the mammalian Bcl-2 superfamilies and to the Caenorhabditis elegans Ced-9. When the two domains, BH1 and BH2, are analyzed separately, again the highest similarity was found to the members of the Bcl-2 superfamily, but a clearly lower relationship to the C. elegans BH1 and BH2 domains in Ced-9. In unrooted phylogenetic trees the sponge BH1 and BH2 are grouped among the mammalian sequences and are only distantly related to the C. elegans BH domains. The analysis of the gene structure of the G. cydonium BHP showed that the single intron present is located within the BH2 domain at the same position as in C. elegans and rat Bcl-x_L. In addition, a sponge molecule comprising two death domains has been characterized from G. cydonium. The two death domains of the potential proapoptotic molecule GC_DD2, M _r 24,970, share a high similarity with the Fas-FADD/MORT1 domains. A death domain-containing molecule has not been identified in the C. elegans genome. The phylogenetic analysis revealed that the sponge domain originated from an ankyrin building block from which the mammalian Fas-FADD/MORT1 evolved. It is suggested that the apoptotic pathways that involve members of the Bcl-2 superfamily and of the death receptors are already present in the lowest metazoan phylum, the Porifera. Received: 27 July 1999 / Accepted: 28 December 1999     

Stabilizing mutations increase secretion of functional soluble TCR-Ig fusion proteins

Background  

Whereas T cell receptors (TCRs) detect peptide/major histocompatibility complexes (pMHCs) with exquisite specificity, there are challenges regarding their expression and use as soluble detection molecules due to molecular instability. We have investigated strategies for the production of TCR-immunoglobulin (Ig) fusion proteins. Two different TCRs that are characteristic of a mouse model for idiotype (Id) dependent immune regulation were engineered. They are structurally unrelated with different variable (V), diversity (D) and joining (J) segments, but each share one V gene segment, either V_α or V_β, with the well characterized murine TCR, 2C.     

神经元GPI锚定蛋白结构与功能研究进展

神经系统中的IgSF蛋白是一类重要的神经细胞表面分子,种类繁多,功能多样,其中一类分子缺乏跨膜区,通过GPI锚结合在细胞膜上,表现出结构和功能的特异性。本文对该类神经元GPI锚定蛋白分子作一系统介绍。根据结构特点,神经元GPI锚定蛋白分子可分为三类,各类分子通过Ig结构域与其自身或其它蛋白分子进行顺式或反式结合,调节神经细胞活动。     

Molecular evolution of immunoglobulin superfamily genes in primates

Genes of the immunoglobulin superfamily (IgSF) have a wide variety of cellular activities. In this study, we investigated molecular evolution of IgSF genes in primates by comparing orthologous sequences of 249 IgSF genes among human, chimpanzee, orangutan, rhesus macaque, and common marmoset. To evaluate the non-synonymous/synonymous substitution ratio (ω), we applied Bn-Bs program and PAML program. IgSF genes were classified into 11 functional categories based on the Gene Ontology (GO) database. Among them, IgSF genes in three functional categories, immune system process (GO:0002376), defense response (GO:0006952), and multi-organism process (GO:0051704), which are tightly linked to the regulation of immune system had much higher values of ω than genes in the other GO categories. In addition, we estimated the average values of ω for each primate lineage. Although each primate lineage had comparable average values of ω, the human lineage showed the lowest ω value for the immune-related genes. Furthermore, 11 IgSF genes, SIGLEC5, SLAMF6, CD33, CD3E, CEACAM8, CD3G, FCER1A, CD48, CD4, TIM4, and FCGR2A, were implied to have been under positive selective pressure during the course of primate evolution. Further sequence analyses of CD3E and CD3G from 23 primate species suggested that the Ig domains of CD3E and CD3G underwent the positive Darwinian selection.     

Neurotractin, a novel neurite outgrowth-promoting Ig-like protein that interacts with CEPU-1 and LAMP.

The formation of axon tracts in nervous system histogenesis is the result of selective axon fasciculation and specific growth cone guidance in embryonic development. One group of proteins implicated in neurite outgrowth, fasciculation, and guidance is the neural members of the Ig superfamily (IgSF). In an attempt to identify and characterize new proteins of this superfamily in the developing nervous system, we used a PCR-based strategy with degenerated primers that represent conserved sequences around the characteristic cysteine residues of Ig-like domains. Using this approach, we identified a novel neural IgSF member, termed neurotractin. This GPI-linked cell surface glycoprotein is composed of three Ig-like domains and belongs to the IgLON subgroup of neural IgSF members. It is expressed in two isoforms with apparent molecular masses of 50 and 37 kD, termed L-form and S-form, respectively. Monoclonal antibodies were used to analyze its biochemical features and histological distribution. Neurotractin is restricted to subsets of developing commissural and longitudinal axon tracts in the chick central nervous system. Recombinant neurotractin promotes neurite outgrowth of telencephalic neurons and interacts with the IgSF members CEPU-1 (KD = 3 x 10(-8) M) and LAMP. Our data suggest that neurotractin participates in the regulation of neurite outgrowth in the developing brain.     

Expression pattern of immunoglobulin superfamily members in the silkworm, Bombyx mori

Immunoglobulin superfamily (IgSF) proteins are involved in cell adhesion, cell communication and immune functions. In this study, 152 IgSF genes containing at least one immunoglobulin (Ig) domain were predicted in the Bombyx mori silkworm genome. Of these, 145 were distributed on 25 chromosomes with no genes on chromosomes 16, 18 and 26. Multiple sequence alignments and phylogenetic evolution analysis indicated that IgSFs evolved rapidly. Gene ontology (GO) annotation indicated that IgSF members functioned as cellular components and in molecular functions and biological processes. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis suggested that IgSF proteins were involved in signal transduction, signaling molecules and interaction, and cell communication. Microarray-based expression data showed tissue expression for 136 genes in anterior silkgland, middle silkgland, posterior silkgland, testis, ovary, fat body, midgut, integument, hemocyte, malpighian tubule and head. Expression pattern of IgSF genes in the silkworm ovary and midgut was analyzed by RNA-Seq. Expression of 105 genes was detected in the ovary in strain Dazao. Expression in the midgut was detected for 74 genes in strain Lan5 and 75 genes in strain Ou17. Expression of 34 IgSF genes in the midgut relative to the actin A3 gene was significantly different between strains Lan5 and Ou17. Furthermore, 1 IgSF gene was upregulated and 1 IgSF gene was downregulated in strain Lan5, and 4 IgSF genes were upregulated and 2 IgSF genes were downregulated in strain Ou17 after silkworms were challenged with B. mori cypovirus (BmCPV), indicating potential involvement in the response to BmCPV-infection. These results provide an overview of IgSF family members in silkworms, and lay the foundation for further functional studies.     

Expanding our understanding of immunoglobulin, T-cell antigen receptor, and novel immune-type receptor genes: a subset of the immunoglobulin gene superfamily

 The immunoglobulin superfamily (IgSF) is an extensively diversified multigene family whose members share a common structural feature, the Ig fold. Members of the Ig/T-cell antigen receptor (TCR) subset of the IgSF mediate antigen-specific recognition in adaptive immune responses. Antigen-binding receptors belonging to this subset are present in all species of jawed vertebrates. To explore whether there are additional structurally related but otherwise distinct members of this subset, we have developed a technique termed the short-primer polymerase chain reaction (PCR) that targets structurally conserved short motifs in the Ig fold. Large-scale sequencing efforts and recent advances in information biotechnology, including "electronic PCR," provide additional computational means to implement similarly directed searches within databases. The use of these approaches has led to the discoveries of Ig/TCR homologues in a variety of phylogenetically diverse organisms, a diversified family of novel immune-type receptor genes, as well as a novel human IgSF member. The potential of random sequencing efforts and virtual screening of databases is described in the context of two novel genes in bony fish. The various methodologies that are discussed and the examples shown provide means for further investigating, and/or elucidating novel, IgSF receptors as well as components of pathways that are involved in immune responses in both traditional and nontraditional model systems.     

Channel catfish leukocyte immune-type receptors contain a putative MHC class I binding site

The recent identification of a large and diverse family of leukocyte immune-type receptors (IpLITRs) in channel catfish (Ictalurus punctatus) indicates that immunoglobulin superfamily (IgSF) members related to both mammalian Fc receptors (FcRs) and leukocyte receptor complex (LRC)-encoded proteins exist in fish. In the present study, it was found that IpLITR messages were preferentially up regulated in catfish peripheral blood leukocytes (PBL) and clonal cytotoxic T cells (CTL) after alloantigen stimulation. Detailed sequence analyses of the expressed IpLITR cDNAs from two clonal CTL lines indicated an unexpectedly large array of putative activatory and inhibitory IpLITR-types containing variable numbers of extracellular immunoglobulin (Ig)-like domains. Importantly, all expressed IpLITRs shared similar membrane distal Ig domains (i.e., D1 and D2), suggesting that they may bind a common type of ligand. Sequence alignments and comparative homology modeling revealed that IpLITR domains, D1 and D2, have similar predicted 3-D structural properties with the corresponding domains of the human LRC-encoded leukocyte Ig-like receptor (LILR) family. Furthermore, conservation of key major histocompatibility class I (MHC I)-binding residues were located at similar positions within the membrane distal tip of D1 between representative IpLITRs and group 1 LILRs. Taken together, these results suggest that fish LITRs have an orthologous relationship to LRC-encoded receptors such as the human LILRs and could potentially function as a diverse family of MHC class I-binding receptors.     

Structure of two FREP genes that combine IgSF and fibrinogen domains, with comments on diversity of the FREP gene family in the snail Biomphalaria glabrata

Upon exposure to infection with digenetic trematodes such as Echinostoma paraensei, the freshwater snail Biomphalaria glabrata produces increased quantities of hemolymph lectins, some of which are unique polypeptides containing both immunoglobulin superfamily (IgSF) and fibrinogen domains. These unusual lectins have been termed fibrinogen-related proteins (FREPs), and recognize and precipitate digenean antigens. We here report 11 distinct FREP-encoding sequences from B. glabrata, and provide the complete genomic sequence for two of the most frequently recovered FREPs. The unique juxtaposition of IgSF and fibrinogen domains, previously known only from incomplete cDNAs, is confirmed. Sequences corresponding to known peptides derived from FREPs from hemolymph were found in one of these genes. Both genes contain four exons, the first encodes a putative signal peptide, the second and third a portion of an IgSF-type loop, and the fourth a fibrinogen domain. Cysteines, postulated to form an intrachain loop, are present in the IgSF domain and are separated from one another by 78 or 79 residues. The IgSF sequences most closely resemble V (variable)-type Ig domains, based on canonical and hydrophobic residues and predicted secondary structure. Some minor differences in genomic fragments isolated for each of the two sequences were noted and may represent allelic variants. The results may be of relevance in understanding the role of B. glabrata in transmission of Schistosoma mansoni, a digenean parasite that infects nearly 100 million people in the tropics.     

Neurite Fasciculation Mediated by Complexes of Axonin-1 and Ng Cell Adhesion Molecule

Neural cell adhesion molecules composed of immunoglobulin and fibronectin type III-like domains have been implicated in cell adhesion, neurite outgrowth, and fasciculation. Axonin-1 and Ng cell adhesion molecule (NgCAM), two molecules with predominantly axonal expression exhibit homophilic interactions across the extracellular space (axonin- 1/axonin-1 and NgCAM/NgCAM) and a heterophilic interaction (axonin-1–NgCAM) that occurs exclusively in the plane of the same membrane (cis-interaction). Using domain deletion mutants we localized the NgCAM homophilic binding in the Ig domains 1-4 whereas heterophilic binding to axonin-1 was localized in the Ig domains 2-4 and the third FnIII domain. The NgCAM–NgCAM interaction could be established simultaneously with the axonin-1–NgCAM interaction. In contrast, the axonin-1–NgCAM interaction excluded axonin-1/axonin-1 binding. These results and the examination of the coclustering of axonin-1 and NgCAM at cell contacts, suggest that intercellular contact is mediated by a symmetric axonin-1₂/NgCAM₂ tetramer, in which homophilic NgCAM binding across the extracellular space occurs simultaneously with a cis-heterophilic interaction of axonin-1 and NgCAM. The enhanced neurite fasciculation after overexpression of NgCAM by adenoviral vectors indicates that NgCAM is the limiting component for the formation of the axonin-1₂/NgCAM₂ complexes and, thus, neurite fasciculation in DRG neurons.     

A novel family of diversified immunoregulatory receptors in teleosts is homologous to both mammalian Fc receptors and molecules encoded within the leukocyte receptor complex

Three novel and closely related leukocyte immune-type receptors (IpLITR) have been identified in channel catfish (Ictalurus punctatus). These receptors belong to a large polymorphic and polygenic subset of the Ig superfamily with members located on at least three independently segregating loci. Like mammalian and avian innate immune regulatory receptors, IpLITRs have both putative inhibitory and stimulatory forms, with multiple types coexpressed in various lymphoid tissues and clonal leukocyte cell lines. IpLITRs have an unusual and novel relationship to mammalian and avian innate immune receptors: the membrane distal Ig domains of an individual IpLITR are related to fragment crystallizable receptors (FcRs) and FcR-like proteins, whereas the membrane proximal Ig domains are related to several leukocyte receptor complex encoded receptors. This unique composition of Ig domains within individual receptors supports the hypothesis that functionally and genomically distinct immune receptor families found in tetrapods may have evolved from such ancestral genes by duplication and recombination events. Furthermore, the discovery of a large heterogeneous family of immunoregulatory receptors in teleosts, reminiscent of amphibian, avian, and mammalian Ig-like receptors, suggests that complex innate immune receptor networks have been conserved during vertebrate evolution.Electronic supplementary material Supplementary material is available for this article at and is accessible for authorized users. GenBank Submissions: The sequences presented in this article have been submitted to GenBank under the following accession numbers: AAW82352, IpLITR1; AAW82353, IpLITR2; AAW82354, IpLITR3.     

Molecular cloning, characterization, and chromosomal localization of the mouse homologue of CD84, a member of the CD2 family of cell surface molecules

 CD84 is a member of the immunoglobulin gene superfamily (IgSF) with two Ig-like domains expressed primarily on B lymphocytes and macrophages. Here we describe the cloning of the mouse homologue of human CD84. Mouse CD84 cDNA clones were isolated from a macrophage library. The nucleotide sequence of mouse CD84 was shown to include an open reading frame encoding a putative 329 amino acid protein composed of a 21 amino acid leader peptide, two extracellular immunoglobulin (Ig)-like domains, a hydrophobic transmembrane region, and an 87 amino acid cytoplasmic domain. Mouse CD84 shares 57.3% amino acid sequence identity (88.7%, considering conservative amino acid substitutions) with the human homologue. Chromosome localization studies mapped the mouse CD84 gene to distal chromosome 1 adjacent to the gene for Ly-9, placing it close to the region where other members of the CD2 IgSF (CD48 and 2B4) have been mapped. Northern blot analysis revealed that the expression of mouse CD84 was predominantly restricted to hematopoietic tissues. Two species of mRNA of 3.6 kilobases (kb) and 1.5 kb were observed. The finding that the pattern of expression was restricted to the hematopoietic system and the conserved sequence of the mouse CD84 homologue suggests that the function of the CD84 glycoprotein may be similar in humans and mice. Received: 1 July 1998 / Revised: 31 August 1998     

Recognition of defined epitopes by affinity‐purified anti‐immunoglobulin Fab autoantibodies isolated from HIV‐infected humans

Infection of humans with HIV‐1 has previously been independently shown to result in the generation of autoantibodies (AAbs) reactive with immunoglobulin Fab fragments (Heidelberg), and with autoantibodies to T‐cell receptors (TCRs) (Tucson). Here, we carry out epitope mapping studies of affinity‐purified AAbs to Fab fragments prepared from individual HIV‐positive patients for their capacity to bind recombinant constructs and peptide‐defined epitopes modeling TCR and Ig light chains. Some affinity‐purified autoantibodies reacted strongly with TCRs expressed by intact T‐cells, and recombinant V_α/V_β constructs as well as with certain synthetic peptide epitopes. The binding reactions of affinity‐purified AAbs of individual patients were distinct, and the AAb preparations consisted of populations of polyclonal lgs as reflected in specificity and isotype. AAb pools from individual patients all bound particular regions of TCR and Ig chains defined by comprehensive peptide synthesis including the CDR1 and Fr3 segments of the variable domains and the joining segment/switch peptide. In addition, other reactivities to restricted regions of α, β and λ light chains were documented. These results substantiate the cross‐reactivity of TCR and Ig–Fab determinants, and are consistent with the hypothesis that autoantibodies arising as a consequence of HIV infection can have an immunomodulatory role. Copyright © 1999 John Wiley & Sons, Ltd.     

Parasite-responsive IgSF members in the snail Biomphalaria glabrata: characterization of novel genes with tandemly arranged IgSF domains and a fibrinogen domain

Two novel genes of the immunoglobulin superfamily (IgSF), FREP3 and FREP7, are reported from the snail Biomphalaria glabrata, a prominent intermediate host of the human parasite Schistosoma mansoni. They resemble other B. glabrata genes that encode fibrinogen-related proteins (FREPs), but differ in that they encode proteins with two tandemly arranged IgSF domains followed by a C-terminal fibrinogen domain. FREPs are hemolymph proteins that increase in abundance following exposure to a digenetic trematode, Echinostoma paraensei, and that bind to and precipitate parasite antigens. Within each gene, the two IgSF-coding regions are dissimilar from one another: the N-terminal IgSF1 domain is encoded by a single exon whereas the downstream IgSF2 domain is encoded by three exons. For both FREPs 3 and 7, the IgSF2 domain belongs to the variable (V) type, whereas the IgSF1 domain is not easily classified with respect to IgSF type. The fibrinogen-encoding region in both genes is relatively conserved and lacks introns. FREP3 exhibits extensive variation in the IgSF1 region. A ratio of nonsynonymous versus synonymous substitutions of 2.56 suggests that this region is under positive selection. A genomic fragment identifiable as FREP7 but lacking an exon was also found, further suggestive of variability within FREP IgSF-encoding regions. Insofar as FREPs are hypothesized to function in nonself recognition, the identification of additional novel FREP genes as part of a growing gene family in B. glabrata is of interest. Such genes, particularly given their variable nature, serve as a model to study the complexity of invertebrate defense responses.