Immune Related Genes Underpin the Evolution of Adaptive Immunity in Jawless Vertebrates

The study of immune related genes in lampreys and hagfish provides a unique perspective on the evolutionary genetic underpinnings of adaptive immunity and the evolution of vertebrate genomes. Separated from their jawed cousins at the stem of the vertebrate lineage, these jawless vertebrates have many of the gene families and gene regulatory networks associated with the defining morphological and physiological features of vertebrates. These include genes vital for innate immunity, inflammation, wound healing, protein degradation, and the development, signaling and trafficking of lymphocytes. Jawless vertebrates recognize antigen by using leucine-rich repeat (LRR) based variable lymphocyte receptors (VLRs), which are very different from the immunoglobulin (Ig) based T cell receptor (TCR) and B cell receptor (BCR) used for antigen recognition by jawed vertebrates. The somatically constructed VLR genes are expressed in monoallelic fashion by T-like and B-like lymphocytes. Jawless and jawed vertebrates thus share many of the genes that provide the molecular infrastructure and physiological context for adaptive immune responses, yet use entirely different genes and mechanisms of combinatorial assembly to generate diverse repertoires of antigen recognition receptors.     

Genome biology of the cyclostomes and insights into the evolutionary biology of vertebrate genomes

The jawless vertebrates (lamprey and hagfish) are the closest extant outgroups to all jawed vertebrates (gnathostomes) and can therefore provide critical insight into the evolution and basic biology of vertebrate genomes. As such, it is notable that the genomes of lamprey and hagfish possess a capacity for rearrangement that is beyond anything known from the gnathostomes. Like the jawed vertebrates, lamprey and hagfish undergo rearrangement of adaptive immune receptors. However, the receptors and the mechanisms for rearrangement that are utilized by jawless vertebrates clearly evolved independently of the gnathostome system. Unlike the jawed vertebrates, lamprey and hagfish also undergo extensive programmed rearrangements of the genome during embryonic development. By considering these fascinating genome biologies in the context of proposed (albeit contentious) phylogenetic relationships among lamprey, hagfish, and gnathostomes, we can begin to understand the evolutionary history of the vertebrate genome. Specifically, the deep shared ancestry and rapid divergence of lampreys, hagfish and gnathostomes is considered evidence that the two versions of programmed rearrangement present in lamprey and hagfish (embryonic and immune receptor) were present in an ancestral lineage that existed more than 400 million years ago and perhaps included the ancestor of the jawed vertebrates. Validating this premise will require better characterization of the genome sequence and mechanisms of rearrangement in lamprey and hagfish.     

无颌类脊椎动物适应性免疫系统的研究进展

在以七鳃鳗和盲鳗为代表的无颌类脊椎动物中, 虽然发现了与有颌类脊椎动物T细胞受体(T-cell receptors, TLRs)、B细胞受体 (B-cell receptors, BCRs)可变区具有相似结构的先天性免疫受体, 却从未发现有颌类脊椎动物适应性免疫系统的核心组分: TCRs、BCRs、组织相容性复合体 (Major histocompatibility complex, MHC)。因此, 长期以来, 人们一直认为适应性免疫系统只存在于有颌类脊椎动物中。但最近的一项发现彻底改变了这一传统观念, 即在无颌类脊椎动物中, 存在一种新型可变淋巴细胞受体VLRs(Variable lymphocyte receptors), VLRs通过改变亮氨酸富集序列LRRs(Leucine-rich repeats)的插入情况, 实现对特异性抗原的高效识别。晶体衍射分析发现, 盲鳗的VLRs呈现一种“马蹄”型结构, 抗原结合位点则位于“马蹄”的凹面区。分泌型的VLRs以四聚体或五聚体的形式识别、结合特异性抗原。综上所述, 无颌类和有颌类脊椎动物应用不同的抗原识别系统完成适应性免疫反应。文章对近年来无颌类脊椎动物适应性免疫系统相关分子的研究进展加以概述, 为揭示适应性免疫系统起源与进化问题提供有益参考。     

Variable domains and a VpreB-like molecule are present in a jawless vertebrate

Immunoglobulins (Igs) and T cell antigen receptors (TCRs) that undergo somatic diversification have not been identified in the two extant orders of jawless vertebrates, which occupy essential positions in terms of understanding the evolution of the emergence of adaptive immunity. Using a single motif-dependent PCR-based approach coupled with a vector that allows selection of cDNAs encoding secretion signal sequences, four different genes encoding Ig V-type domains were identified in the sea lamprey (Petromyzon marinus). One of the predicted proteins encoded by these genes shares structural characteristics with mammalian VpreB molecules, including the absence of a recognizable transmembrane region, a relatively high proportion of charged amino acids in its C-terminal tail and distinctive features of its secretion signal peptide. This is the first indication of a molecule related to the B cell receptor (BCR) complex in a species that diverged prior to the jawed vertebrates in which RAG-mediated adaptive immunity is first encountered.Sequences described in this paper have been deposited in GenBank, with accession numbers AY576797–AY576800.     

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.     

Hagfish leukocytes express a paired receptor family with a variable domain resembling those of antigen receptors

Jawed vertebrates are equipped with TCR and BCR with the capacity to rearrange their V domains. By contrast, jawless vertebrates, represented by hagfish and lampreys, apparently lack such receptors. We describe in this study a family of hagfish genes carrying a single V-type domain resembling those of TCR/BCR. This multigene family, which we call agnathan paired receptors resembling Ag receptors (APAR), is expressed in leukocytes and predicted to encode a group of membrane glycoproteins with organizations characteristic of paired Ig-like receptors, consisting of activating and inhibitory forms. APAR has a J region in its V-type domain, and its V and J regions are encoded in a single exon. Thus, APAR is a member of the emerging families of diversified, innate immune-type receptors with TCR/BCR-like V-type domains and has many of the features expected for a primordial TCR/BCR-like receptor. The extracellular domain of APAR may be descended from a V-type domain postulated to have acquired recombination signal sequences in a jawed vertebrate lineage.     

The evolution of adaptive immune systems

A clonally diverse anticipatory repertoire in which each lymphocyte bears a unique antigen receptor is the central feature of the adaptive immune system that evolved in our vertebrate ancestors. The survival advantage gained through adding this type of adaptive immune system to a pre-existing innate immune system led to the evolution of alternative ways for lymphocytes to generate diverse antigen receptors for use in recognizing and repelling pathogen invaders. All jawed vertebrates assemble their antigen-receptor genes through recombinatorial rearrangement of different immunoglobulin or T cell receptor gene segments. The surviving jawless vertebrates, lampreys and hagfish, instead solved the receptor diversification problem by the recombinatorial assembly of leucine-rich-repeat genetic modules to encode variable lymphocyte receptors. The convergent evolution of these remarkably different adaptive immune systems involved innovative genetic modification of innate-immune-system components.     

Anatomical and Physiological Peculiarities of the Heart in Jawless and Jawed Fish

Journal of Evolutionary Biochemistry and Physiology - The heart of jawless fish (Cyclostomata; lamprey, hagfish) and jawed fish (Teleostei) is homologous to the heart of higher vertebrates. A study...     

Design principles of adaptive immune systems

Both jawless vertebrates, such as lampreys and hagfish, and jawed vertebrates (encompassing species as diverse as sharks and humans) have an adaptive immune system that is based on somatically diversified and clonally expressed antigen receptors. Although the molecular nature of the antigen receptors and the mechanisms of their assembly are different, recent findings suggest that the general design principles underlying the two adaptive immune systems are surprisingly similar. The identification of such commonalities promises to further our understanding of the mammalian immune system and to inspire the development of new strategies for medical interventions targeting the consequences of faulty immune functions.     

The origin of developmental mechanisms underlying vertebral elements: implications from hagfish evo-devo

The origins of the vertebral elements and the underlying developmental mechanisms have so far remained unclear, largely due to the unusual axial skeletal morphology of hagfish, one of two extant jawless vertebrate clades. Hagfish axial supporting tissue is generally believed to consist of the notochord and cartilaginous fin rays only. However, careful investigations of whether vertebral elements are truly absent in hagfish are scarce, and it is also unclear whether the axial skeletal morphology of the hagfish is an ancestral or a derived condition. To address these questions, we re-examined the axial skeletal morphology of the Japanese inshore hagfish (Eptatretus burgeri). Based on a report published a century ago which implied the existence of vertebral elements in hagfish, we conducted anatomical and histological analyses of the hagfish axial skeletal systems and their development. Through this analysis, we demonstrate that hagfish possesses sclerotome-derived cartilaginous vertebral elements at the ventral aspect of the notochord. Based on (i) molecular phylogenetic evidence in support of the monophyly of cyclostomes (hagfish and lampreys) and jawed vertebrates (gnathostomes), and (ii) the morphology of the vertebral elements in extant gnathostomes and cyclostomes, we propose that the embryos of the common ancestor of all vertebrates would have possessed sclerotomal cells that formed the segmentally arranged vertebral elements attached to the notochord. We also conclude that the underlying developmental mechanisms are likely to have been conserved among extinct jawless vertebrates and modern gnathostomes.     

Members of the Ikaros gene family are present in early representative vertebrates

Members of the Ikaros multigene family of zinc finger proteins are expressed in a tissue-specific manner and most are critical determinants in the development of both the B and T lymphocytes as well as NK and dendritic APC lineages. A PCR amplification strategy that is based on regions of shared sequence identity in Ikaros multigene family members found in mammals and several other vertebrates has led to the recovery of cDNAs that represent the orthologues of Ikaros, Aiolos, Helios, and Eos in Raja eglanteria (clearnose skate), a cartilaginous fish that is representative of an early divergence event in the phylogenetic diversification of the vertebrates. The tissue-specific patterns of expression for at least two of the four Ikaros family members in skate resemble the patterns observed in mammals, i.e., in hematopoietic tissues. Prominent expression of Ikaros in skate also is found in the lymphoid Leydig organ and epigonal tissues, which are unique to cartilaginous fish. An Ikaros-related gene has been identified in Petromyzon marinus (sea lamprey), a jawless vertebrate species, in which neither Ig nor TCRs have been identified. In addition to establishing a high degree of evolutionary conservation of the Ikaros multigene family from cartilaginous fish through mammals, these studies define a possible link between factors that regulate the differentiation of immune-type cells in the jawed vertebrates and related factors of unknown function in jawless vertebrates.     

Evolutionary crossroads in developmental biology: cyclostomes (lamprey and hagfish)

Lampreys and hagfish, which together are known as the cyclostomes or 'agnathans', are the only surviving lineages of jawless fish. They diverged early in vertebrate evolution, before the origin of the hinged jaws that are characteristic of gnathostome (jawed) vertebrates and before the evolution of paired appendages. However, they do share numerous characteristics with jawed vertebrates. Studies of cyclostome development can thus help us to understand when, and how, key aspects of the vertebrate body evolved. Here, we summarise the development of cyclostomes, highlighting the key species studied and experimental methods available. We then discuss how studies of cyclostomes have provided important insight into the evolution of fins, jaws, skeleton and neural crest.     

The leukocyte common antigen (CD45) of the Pacific hagfish,Eptatretus stoutii: implications for the primordial function of CD45

CD45, originally known as the leukocyte common antigen, is a prototypical transmembrane protein tyrosine phosphatase that plays a critical role in signal transduction through T-cell and B-cell receptors, as well as in T-cell and B-cell development. In the present study, we show that the Pacific hagfish, widely believed to lack the adaptive immune system, has CD45. The presence of CD45 in jawless fish is consistent with the recent discovery that CD45 also plays a crucial role in innate immunity via the regulation of signaling through type I and type II cytokine receptors. It is likely that CD45 was recruited to activate lymphocytes through antigen receptors encoded by rearranging genes in jawed vertebrates.     

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     

The mitochondrial DNA molecule of the hagfish (myxine glutinosa) and vertebrate phylogeny

The vertebrates are traditionally classified into two distinct groups, Agnatha (jawless vertebrates) and Gnathostomata (jawed vertebrates). Extant agnathans are represented by hagfishes (Myxiniformes) and lampreys (Petromyzontiformes), frequently grouped together within the Cyclostomata. Whereas the recognition of the Gnathostomata as a clade is commonly acknowledged, a consensus has not been reached regarding whether or not Cyclostomata represents a clade. In the present study we have used newly established sequences of the protein-coding genes of the mitochondrial DNA molecule of the hagfish to explore agnathan and gnathostome relationships. The phylogenetic analysis of Pisces, using echinoderms as outgroup, placed the hagfish as a sister group of Vertebrata sensu stricto, i.e., the lamprey and the gnathostomes. The phylogenetic analysis of the Gnathostomata identified a basal divergence between gnathostome fishes and a branch leading to birds and mammals, i.e., between ``Anamnia' and Amniota. The lungfish has a basal position among gnathostome fishes with the teleosts as the most recently evolving lineage. The findings portray a hitherto unrecognized polarity in the evolution of bony fishes. The presently established relationships are incompatible with previous molecular studies. Received: 15 August 1997 / Accepted: 1 October 1997     

Two variable lymphocyte receptor genes of the inshore hagfish are located far apart on the same chromosome

Variable lymphocyte receptors (VLR) generate enormous diversity through assembling highly diverse leucine-rich repeat (LRR) modules and presumably function as antigen receptors in jawless vertebrates. The hagfish, which constitute major extant members of jawless vertebrates along with lampreys, have two VLR genes designated VLRA and VLRB, whereas only a single VLR gene has been identified in the lamprey. In the present study, we show by fluorescence in situ hybridization (FISH) that hagfish VLRA and VLRB are located on the same chromosome, but are far apart from each other. Analysis of available inshore hagfish complementary DNA sequences indicates that VLRA and VLRB do not share a LRR module with an identical nucleotide sequence. Physical separation of VLRA and VLRB is consistent with this observation and indicates that the two VLR genes function as separate units. The FISH protocol developed in this study should be useful for the analysis of the agnathan genome. J. K. is a research fellow of the Japan Society for the Promotion of Science.