Lampetra Japonica Research Articles

Evolution and expression of TLR5a and TLR5b in lamprey (Lampetra japonica)

The lamprey serves as a key model organism for studying the origin and evolution of species, embryonic development, and the immune system. The immune system primarily relies on pattern recognition receptors, including Toll-like receptors (TLRs), with Toll-like receptor 5 (TLR5) having a particularly complex evolutionary history. Currently, although TLR5 is being identified in an expanding array of taxonomic groups, a comprehensive study on its evolutionary aspects is yet to be conducted. In this study, we identified Lj-TLR5a and Lj-TLR5b in Lampetra japonica and examined their distribution across various tissues in this species. Furthermore, we conducted preliminary investigations into their immune functions and discovered that, as primitive genes, they are highly sensitive to various pathogens. Upon recognizing flagellar proteins, both Lj-TLR5a and Lj-TLR5b work together; however, these TLRs may function independently in response to other stimuli. Subsequently, we performed comprehensive structural and evolutionary analyses of the TLR5 family, incorporating TLR5 data from various species at different evolutionary stages. Our findings revealed that TLR5a and TLR5b in lamprey are in a relatively primitive evolutionary state. Meanwhile, TLR5L differentiated during the early stages of evolution and exhibits a pseudogenic trend throughout this evolutionary process; notably, this TLR is currently preserved only in certain amphibian and reptile species. In cartilaginous fish, only one type of TLR5 is usually retained, whereas bony fish typically possess both TLR5 and TLR5S. TLR5S has a relatively simple structure, likely arising from repetitive whole-genome events in bony fishes. Among bony fishes, flesh-finned fish were found to retain only one TLR5, which eventually evolved into the TLR5 found in quadrupeds. In summary, this study provides significant insights into the origin and evolution of the TLR5 family by analyzing the evolutionary status and immune functions of Lj-TLR5a and Lj-TLR5b in Japanese lampreys.

Form-assortative mating behaviors of individuals from parasitic and non-parasitic populations of Arctic lamprey (Lethenteron camtschaticum)

Abstract Evolutionary theory predicts that assortative mating is crucial for sympatric speciation by generating reproductive isolation between diverging populations. Here, we investigate the potential of form-assortative mating, an assumed mating pattern in lampreys, for sympatric speciation. By continuously recording mating activity between anadromous and freshwater-resident forms of L. camtschaticum that greatly differ in body size, we show that lampreys tend to mate with individuals of similar size in experimental conditions. However, we highlight that this pattern does not result from a choice of same-form partner but is the result of the simultaneous action of a preference of males – whatever their size – for large anadromous females, a higher competitive ability of aggressive males and physical constraints on heteroform pairs. Moreover, we do not advocate that sympatric speciation, as the sole consequence of form-assortative mating through sexual selection, is a plausible mechanism for the diversification of lampreys as a significant number of sneaking behaviors were observed in freshwater-resident males toward large anadromous females. Broader attention should be given to mechanisms other than sexual selection that may lead to form-assortative mating, such as variations in spatial or temporal distribution of alternative forms during reproductive season.

Bactericidal ability of target acidic phospholipids and phagocytosis of CDC42 GTPase-mediated cytoskeletal rearrangement underlie functional conservation of CXCL12 in vertebrates.

Chemokine CXCL12 plays a crucial role in both direct bactericidal activity and phagocytosis in humans. However, the mechanisms and evolutionary functions of these processes in vertebrates remain largely unknown. In this study, we found that the direct bactericidal activity of CXCL12 is highly conserved across various vertebrate lineages, including Arctic lamprey (Lampetra japonica), Basking shark (Cetorhinus maximus), grass carp (Ctenopharyngodon idella), Western clawed frog (Xenopus tropicalis), Green anole (Anolis carolinensis), chicken (Gallus gallus), and human (Homo sapiens). CXCL12 also has been shown to promote phagocytosis in lower and higher vertebrates. We then employed C. idella CXCL12a (CiCXCL12a) as a model to further investigate its immune functions and underlying mechanisms. CiCXCL12a exerts direct broad-spectrum antibacterial activity by targeting bacterial acidic phospholipids, resulting in bacterial cell membrane perforation, and eventual lysis. Monocytes/macrophages are attracted to the infection sites for phagocytosis through the rapid production of CiCXCL12a during bacterial infection. CiCXCL12a induces CDC42 and CDC42 GTPase activation, which in turn mediates F-actin polymerization and cytoskeletal rearrangement. The interaction between F-actin and Aeromonas hydrophila facilitates bacterial internalization into monocytes/macrophages. Additionally, A. hydrophila is colocalized within early endosomes, late endosomes and lysosomes, ultimately degrading within phagolysosomes. CiCXCL12a also activates PI3K-AKT, JAK-STAT5 and MAPK-ERK signaling pathways. Notably, only the PI3K-AKT signaling pathway inhibits LPS-induced monocyte/macrophage apoptosis. Thus, CiCXCL12a plays key roles in reducing tissue bacterial loads, attenuating organ injury, and decreasing mortality rates. Altogether, our findings elucidate the conserved mechanisms underlying CXCL12-mediated bactericidal activity and phagocytosis, providing novel perspectives into the immune functions of CXCL12 in vertebrates.

Variability and phenotypic diversity of the Diplostomum Petromyzifluviatilis Diesing, 1850 trematodes, parasites of lampreys (Lampetrinae, Petromyzontidae)

Variability and phenotypic diversity of Diplostomum petromyzifluviatilis metacercariae from two lamprey species, the river lamprey Lampetra fluviatilis (Baltic Sea basin) and the Pacific lamprey Lethenteron camtschaticum (Arctic Ocean basin), were studied. It was found that metacercariae from both hosts are similar as in the number of variations in body shape, oral and ventral suckers, and pharynx, so in the frequency of occurrence. Differences in population diversity by variations in the shape of the Holdfast organ were found. It was shown that gostal forms possess the similar range of variability of plastic characters, but has different distribution of frequency classes of the variation curve. We hypothesized that the change in the distribution and character of variation in the size of metacercariae from the Arctic lamprey is one of the population mechanisms of species adaptation to the northern limit of its range.

Identification of antibacterial activity of liver-expressed antimicrobial peptide 2 (LEAP2) from primitive vertebrate lamprey

Liver-expressed antimicrobial peptide 2 (LEAP2) is a member of the antimicrobial peptides family and plays a key role in the innate immune system of organisms. LEAP2 orthologs have been identified from a variety of fish species, however, its function in primitive vertebrates has not been clarified. In this study, we cloned and identified Lc-LEAP2 from the primitive jawless vertebrate lamprey (Lethenteron camtschaticum) which includes a 25 amino acids signal peptide and a mature peptide of 47 amino acids. Although sequence similarity was low compared to other species, the mature Lc-LEAP2 possesses four conserved cysteine residues, forming a core structure with two disulfide bonds between the cysteine residues in the relative 1–3 (Cys 58 and Cys 69) and 2–4 (Cys 64 and Cys 74) positions. Lc-LEAP2 was most abundantly expressed in the muscle, supraneural body and buccal gland of lamprey, and was significantly upregulated during LPS and Poly I:C stimulations. The mature peptide was synthesized and characterized for its antibacterial activity against different bacteria. Lc-LEAP2 possessed inhibition of a wide range of bacteria with a dose-dependence, disrupting the integrity of bacterial cell membranes and binding to bacterial genomic DNA, although its inhibitory function is weak compared to that of higher vertebrates. These data suggest that Lc-LEAP2 plays an important role in the innate immunity of lamprey and is of great value in improving resistance to pathogens. In addition, the antimicrobial mechanism of LEAP2 has been highly conserved since its emergence in primitive vertebrates.

Ultrastructure of the lamprey head mesoderm reveals evolution of the vertebrate head

The cranial muscle is a critical component in the vertebrate head for a predatory lifestyle. However, its evolutionary origin and possible segmental nature during embryogenesis have been controversial. In jawed vertebrates, the presence of pre-otic segments similar to trunk somites has been claimed based on developmental observations. However, evaluating such arguments has been hampered by the paucity of research on jawless vertebrates. Here, we discovered different cellular arrangements in the head mesoderm in lamprey embryos (Lethenteron camtschaticum) using serial block-face scanning electron and laser scanning microscopies. These cell populations were morphologically and molecularly different from somites. Furthermore, genetic comparison among deuterostomes revealed that mesodermal gene expression domains were segregated antero-posteriorly in vertebrates, whereas such segregation was not recognized in invertebrate deuterostome embryos. These findings indicate that the vertebrate head mesoderm evolved from the anteroposterior repatterning of an ancient mesoderm and developmentally diversified before the split of jawless and jawed vertebrates.

Evaluating the viability of the use of T‐bar and radiotelemetry tags on prespawn Arctic Lampreys

AbstractObjectiveArctic Lampreys Lethenteron camtschaticum are harvested by subsistence and commercial fisheries in the Yukon–Kuskokwim River region of Alaska; however, baseline population data are deficient. For mark–recapture and telemetry studies to be effectively implemented to assess Arctic Lamprey abundance, migratory and dispersal patterns, and spawning locations, tags must not impact fish survival, physiology, or behavior.MethodsFor this laboratory evaluation, we examined survival, tag retention, swim endurance at short‐term (1 day posttreatment) and long‐term (43 days [~6 weeks] posttreatment) intervals, and incision healing for T‐bar‐tagged and radio‐tagged prespawn Yukon River Arctic Lampreys (N = 216). Six treatment groups were evaluated: control; sham surgery; external T‐bar anchor tag; and small (0.30 g; 0.1–0.4% tag burden [ratio of tag weight to fish body weight]), medium (0.57 g; 0.2–0.8% tag burden), and large (1.50 g; 0.6–1.9% tag burden) internal dummy radio transmitter tags.ResultAlthough all lampreys survived tagging and surgical procedures through the first 4 weeks of the experiment, a higher tag burden was associated with an increased mortality hazard up to 35 weeks postsurgery. Over the 14‐week experimental period, one T‐bar tag and one small radio tag were shed by lampreys. Although treatment was not a significant predictor of swim endurance, a higher tag burden led to reductions in swim endurance at 1 day posttreatment but not at 43 days posttreatment. Healing did not differ among surgical treatment groups, but persistent inflammation was observed at incision sites and skin erosion was observed at antenna protrusion locations.ConclusionThese results indicate that T‐bar anchor tags and internal radio tags are acceptable for Arctic Lamprey tagging studies if the radio tag burden is ≤1.3% for short‐term (≤14‐week) studies and ≤0.5% for long‐term (≥14‐week) studies.

Identification and characterization of tryptophan-kynurenine pathway-related genes involving lamprey (Lampetra japonica) innate immunity.

The tryptophan-kynurenine (TRP-KYN) pathway is involved in several biological functions, including immunosuppression, inflammatory response, and tumor suppression. Six TRP-KYN pathway-related genes, tryptophan 2,3-dioxygenase (TDO), indoleamine 2,3-dioxygenase 2 (IDO2), aminoadipate aminotransferase (AADAT), glutamate oxaloacetate transaminase 2 (GOT2), kynurenine monooxygenase (KMO), and kynureninase (KYNU) have been identified and cloned from the jawless vertebrate lamprey (Lampetra japonica) to gain insights into their evolution and characterization. Expression distribution showed that the key gene Lj-TDO was highly expressed in the oral gland. Real-time quantitative PCR showed that TRP-KYN pathway-related genes were significantly overexpressed after multi-stimulation. RNA interference showed that Lj-IDO2 knockdown regulated the expression of inflammatory factors. In conclusion, our study successfully clarified the ancestral features and functions of the TRP-KYN pathway, while providing valuable insights into the involvement of this pathway in the immune responses of a jawless vertebrate.

Ecotoxicology of mercury concentrations in arctic lamprey (Lethenteron camtschaticum)

Arctic lamprey (Lethenteron camtschaticum) is an important dietary resource for rural and indigenous communities in parts of Alaska, with some commercial use. As with many fish species harvested for human consumption, there are concerns regarding mercury concentrations ([Hg]) in Arctic lamprey that may impact human health. To date, information regarding the life cycle and diet of Arctic lamprey is scarce, with no published studies examining [Hg] in Arctic lamprey tissues. Our goals were to investigate the feeding ecology of Arctic lamprey from the Bering Sea, determine how diet and potential dietary shifts might influence [Hg] in muscle, and determine if current [Hg] may pose a human health risk. The mean total [Hg] in Arctic lamprey muscle (n = 98) was 19 ng/g wet-weight. Log transformed total [Hg] were not associated with any measured biological variables including length, mass, δ13C values, or δ15N values. A stable isotope mixing model estimated that capelin (Mallotus villosus) accounted for 40.0 ± 4.0% of the Arctic lamprey diet, while Pacific sand lance (Ammodytes hexapterus) and Pacific herring (Clupea pallasii) accounted for 37.8 ± 3.1% and 22.2 ± 3.5% respectively. Finally, diet percentage compositions shifted based on size class (i.e., medium versus large). These results indicated that feeding location, bioaccumulation, and biomagnification are not important drivers of [Hg] in Arctic lamprey and current [Hg] do not pose a human health risk. Taken together, this research further expands our knowledge of Arctic lamprey trophic ecology in the eastern Bering Sea.

A picogram BA-ELISA quantification assay for rLj-RGD3, a platelet fibrinogen receptor antagonist, in the rat plasma and its application to a pharmacokinetic study.

rLj-RGD3, a new member of the RGD (Arginine-Glycine-Aspartate)-motif toxin protein family obtained from Lampetra japonica by means of recombinant DNA techniques, has been demonstrated to be a platelet fibrinogen receptor antagonist and holds potential as a drug candidate for a specific indication. The present article reports an innovative validated highly sensitive and specific biotin-avidin enzyme linked immunosorbent assay (BA-ELISA) to provide a bio-analytical method for pharmacokinetic (PK) studies of rLj-RGD3. The concentration of picogram level rLj-RGD3 in rat plasma was measured using the developed double sandwich BA-ELISA assay, which used two mouse anti-rLj-RGD3 monoclonal antibodies that recognize different epitopes for capture and detection. This method was verified to be highly specific (blank plasma did not interfere with detection), precise (RSD <15%), and accurate (86%-113%). Absolute recovery was in the 94%-119% range. The calibration curve showed good linearity within the 50 to 1600 pg/mL range. The LOQ was as low as 50 pg/mL. The above validated assay was successfully employed to assess PK of rLj-RGD3 in rats. After i.v. and s.c. dosing with 30 μg/kg, the rLj-RGD3 plasma concentration declined bi-exponentially with time. This decay was best fitted to a two-compartment model. In conclusion, the BA-ELISA method described here meets all requirements for PK studies of rLj-RGD3 with an effective pharmacological dose in the μg/kg BW range.

A complete prostaglandin pathway from synthesis to inactivation in the oral gland of the jawless vertebrate lamprey, Lethenteron camtschaticum

Information on the prostaglandin pathway in lampreys is limited. Here, five genes related to the prostaglandin pathway from synthesis to inactivation, namely, phospholipase A2, cyclooxygenase-2, prostaglandin E synthase 3, prostaglandin D synthase, and 15-hydroxyprostaglandin dehydrogenase [NAD(+)], were screened and cloned from the lamprey, Lethenteron camtschaticum. Bioinformatic analysis showed that these lamprey genes are relatively conserved with teleost genes in domains, motifs, gene structure and 3D structure. Analysis of expression distribution of the genes in lamprey tissues revealed that a complete prostaglandin pathway from synthesis to inactivation exists in the oral gland of lamprey, especially the key gene of prostaglandin synthesis cyclooxygenase-2, which was highly expressed in the oral gland. Furthermore, cyclooxygenase-2 expression increased after LPS and Poly I:C stimulations. Using our established spatial metabolite database LampreyDB, six prostaglandin-related metabolites were screened from the oral gland of lamprey, four of which were highly expressed in the oral gland. This study provides new insights into prostaglandin synthesis and inactivation pathways in lamprey, thereby improving our understanding of the origin and evolution of the prostaglandin pathway and contributing to the recognition of lamprey regulatory mechanisms in development and immunity.

Peptidomics Analysis Reveals the Buccal Gland of Jawless Vertebrate Lamprey as a Source of Multiple Bioactive Peptides.

Various proteins with antibacterial, anticoagulant, and anti-inflammatory properties have been identified in the buccal glands of jawless blood-sucking vertebrate lampreys. However, studies on endogenous peptides in the buccal gland of lampreys are limited. In this study, 4528 endogenous peptides were identified from 1224 precursor proteins using peptidomics and screened for bioactivity in the buccal glands of the lamprey, Lethenteron camtschaticum. We synthesized four candidate bioactive peptides (VSLNLPYSVVRGEQFVVQA, DIPVPEVPILE, VVQLPPVVLGTFG, and VPPPPLVLPPASVK), calculated their secondary structures, and validated their bioactivity. The results showed that the peptide VSLNLPYSVVRGEQFVVQA possessed anti-inflammatory activity, which significantly increased the expression of anti-inflammatory factors and decreased the expression of inflammatory factors in THP-1 cells. The peptide VVQLPPVVLGTFG showed antibacterial activity against some gram-positive bacteria. The peptide VSLNLPYSVVRGEQFVQA possessed good ACE inhibitory activity at low concentrations, but no dose-related correlation was observed. Our study revealed that the buccal glands of the jawless vertebrate lamprey are a source of multiple bioactive peptides, which will provide new insights into the blood-sucking mechanism of lamprey.

Phylogeny and expression of ADAM10 and ADAM17 homologs in lamprey

The ADAMs (a disintegrin and metalloproteinase) play regulatory roles in cell adhesion, migration and proteolysis. To explore the origin and evolution of ADAMs, this study identified the homologs of adam10 and adam17 in Lampetra morii and Lampetra japonica. Sequence analysis revealed that they share the same genomic structures with their counterparts in jawed vertebrates. The putative proteins possess conserved motifs, including a furin cut site (RXXR) for precursor processing, an enzyme catalytic motif (HEXGEHXXGXXH) for hydrolysis, and a Ca2+-binding motif (CGNXXXEXGEXCD) for stabilizing protein structure. In addition, a substrate recognition domain is present at the membrane-proximal region of lamprey ADAM17. The cytoplasmic region of lamprey ADAM10 contains a potential threonine phosphorylation site which has been shown to be activated by protein kinase C (PKC) in mammals. Both the adam10 and adam17 genes were constitutively expressed in the brain, kidney, and gills and were differentially regulated in the primary blood leukocytes by lipopolysaccharide (LPS) and pokeweed mitogen (PWM). Adam10 was induced by LPS but not PWM; conversely, adam17 was induced by PWM but not LPS. Taken together, our results suggest that the activation pathways and functions of ADAM10 and ADAM17 are conserved in agnathans.

Phylogenetic and functional properties of hagfish neurohypophysial hormone receptors distinct from their jawed vertebrate counterparts

Vertebrate neurohypophysial hormones, i.e., vasopressin- and oxytocin-family peptides, exert versatile physiological actions via distinct G protein-coupled receptors. The neurohypophysial hormone receptor (NHR) family was classically categorized into four subtypes (V1aR, V1bR, V2R and OTR), while recent studies have identified seven subtypes (V1aR, V1bR, V2aR, V2bR, V2cR, V2dR and OTR; V2aR corresponds to the conventional V2R). The vertebrate NHR family were diversified via multiple gene duplication events at different scales. Despite intensive research effort in non-osteichthyes vertebrates such as cartilaginous fish and lamprey, the molecular phylogeny of the NHR family has not been fully understood. In the present study, we focused on the inshore hagfish (Eptatretus burgeri), another group of cyclostomes, and Arctic lamprey (Lethenteron camtschaticum) for comparison. Two putative NHR homologs, which were previously identified only in silico, were cloned from the hagfish and designated as ebV1R and ebV2R. In vitro, ebV1R, as well as two out of five Arctic lamprey NHRs, increased intracellular Ca2+ in response to exogenous neurohypophysial hormones. None of the examined cyclostome NHRs altered intracellular cAMP levels. Transcripts of ebV1R were detected in multiple tissues including the brain and gill, with intense hybridization signals in the hypothalamus and adenohypophysis, while ebV2R was predominantly expressed in the systemic heart. Similarly, Arctic lamprey NHRs showed distinct expression patterns, underscoring the multifunctionality of VT in the cyclostomes as in the gnathostomes. These results and exhaustive gene synteny comparisons provide new insights into the molecular and functional evolution of the neurohypophysial hormone system in vertebrates.

Organization of the body wall in lampreys informs the evolution of the vertebrate paired appendages.

Vertebrate paired appendages are one of the most important evolutionary novelties in vertebrates. During embryogenesis, the skeletal elements of paired appendages differentiate from the somatic mesoderm, which is a layer of lateral plate mesoderm. However, the presence of the somatic mesoderm in the common ancestor of vertebrates has been controversial. To address this problem, it is necessary but insufficient to understand the developmental process of lateral plate mesoderm formation in lamprey (jawless vertebrates) embryos. Here, I show the presence of the somatic mesoderm in lamprey (Lethenteron camtschaticum) embryos using plastic sectioning and transmission electron microscopy analysis. During the early pharyngeal stages, the somatic mesoderm transforms from the lateral plate mesoderm in the trunk region. Soon after, when the cardiac structures were morphologically distinct, the somatic mesoderm was recognized through the cardiac to more caudal regions. These findings indicated that the somatic mesoderm evolved before the emergence of paired appendages. I also discuss the developmental changes in the body wall organization in the common ancestor of vertebrates, which is likely related to the evolution of the paired appendages.

No strict host specificity: Brain metacercariae Diplostomum petromyzifluviatilis Müller (Diesing, 1850) are conspecific with Diplostomum sp. Lineage 4 of Blasco-Costa et al. (2014).

Metacercariae of Diplostomum petromyzifluviatilis (Digenea, Diplostomidae) from the brain of European river lamprey Lampetra fluviatilis from the Baltic Sea basin and Arctic lamprey Lethenteron camtschaticum from the White Sea basin were studied with the use of genetic and morphological methods. Phylogenetic analysis based on cox1 marker showed that the parasites of both lamprey species were conspecific with Diplostomum sp. Lineage 4 of Blasco-Costa et al. (2014). The name Diplostomum petromyzifluviatilis Müller (Diesing, 1850) has historical precedence as a species described from the brain of lampreys and should be used in genus nomenclature. There were no morphological qualitative differences between the metacercariae from the two lamprey species but those from L. fluviatilis were larger than those from L. camtschaticum. We expanded the data on the second intermediate hosts and the localization of D. petromyzifluviatilis, showing that its metacercariae occur not only in the brain of lampreys but also in the brain and the retina of three-spined stickleback Gasterosteus aculeatus and the vitreous humour of the perch Perca fluviatilis across the European part of the Palearctic.

Molecular Characterization of a B Cell Adaptor for Phosphoinositide 3-Kinase Homolog in Lamprey (Lampetra japonica) and Its Function in the Immune Response.

Human B cell adaptor for phosphoinositide 3-kinase (BCAP) is identified as an adaptor protein expressed in B cells and plays a critical immunomodulatory role in B cell receptor signaling and humoral immune response. In the current study, a homolog of BCAP (Lja-BCAP) was identified in Lampetra japonica. The open reading frame of Lja-BCAP contains 2181bp nucleotides and encodes a protein of 726 amino acids. After being stimulated by mixed bacteria, the mRNA and protein expression levels of Lja-BCAP and the activation levels of tyrosine kinases increased significantly in peripheral blood lymphocytes, gills and supraneural myeloid bodies, respectively. However, after the knockdown of Lja-BCAP by RNAi in vivo, the activation of tyrosine kinases was inhibited in the above tissues, which indicated that Lja-BCAP participated in the anti-bacterial immune response of lampreys. After lipopolysaccharide (LPS) stimulation, the expression of Lja-BCAP in peripheral blood lymphocytes, gills and supraneural myeloid bodies were significantly up-regulated 2.5, 2.2, and 11.1 times (p &lt; 0.05) compared to the control group, respectively; while after phytohemagglutinin (PHA) stimulation, the up-regulation of Lja-BCAP was only detected in peripheral blood lymphocytes. The above results show that Lja-BCAP mainly participates in the LPS-mediated immune response of lampreys.

New Insights Into the Lineage-Specific Expansion and Functional Diversification of Lamprey AID/APOBEC Family.

The AID/APOBEC family which converts cytidine to uridine on RNA or DNA experienced dynamic expansion in primates in order to resist exogenous viruses and endogenous retrotransposons. Recently, expansion of AID/APOBEC-like homologs has also been observed in the extant jawless vertebrate lamprey. To reveal what causes such expansion and leads to the functional diversification of lamprey cytosine deaminases (CDAs), we reassessed the CDA genes in Lethenteron japonicum (Lj). We first confirmed the expansion of LjCDA1L1 (CDA1-like 1) genes and found the expression correlation of LjCDA2 and LjCDA1L2 with LjVLRs (variable lymphocyte receptors). Among up to 14 LjCDA1L1 proteins, LjCDA1L1_4a has an extremely high deamination activity on ssDNA and buDNA and, unexpectedly, on dsDNA. LjCDA1L1s can also restrict the infection of HSV-1 particles. Thus, the arms race between the host and pathogens along with the recruitment by VLR assembly may participate together to form a driving force in the expansion and diversification of the lamprey AID/APOBEC family.

Molecular Evolution of Transforming Growth Factor-β (TGF-β) Gene Family and the Functional Characterization of Lamprey TGF-β2.

The transforming growth factor-βs (TGF-βs) are multifunctional cytokines capable of regulating a wide range of cellular behaviors and play a key role in maintaining the homeostasis of the immune system. The TGF-β subfamily, which is only present in deuterostomes, expands from a single gene in invertebrates to multiple members in jawed vertebrates. However, the evolutionary processes of the TGF-β subfamily in vertebrates still lack sufficient elucidation. In this study, the TGF-β homologs are identified at the genome-wide level in the reissner lamprey (Lethenteron reissneri), the sea lamprey (Petromyzon marinus), and the Japanese lamprey (Lampetra japonica), which are the extant representatives of jawless vertebrates with a history of more than 350 million years. The molecular evolutionary analyses reveal that the lamprey TGF-β subfamily contains two members representing ancestors of TGF-β2 and 3 in vertebrates, respectively, but TGF-β1 is absent. The transcriptional expression patterns show that the lamprey TGF-β2 may play a central regulatory role in the innate immune response of the lamprey since it exhibits a more rapid and significant upregulation of expression than TGF-β3 during lipopolysaccharide stimuli. The incorporation of BrdU assay reveals that the lamprey TGF-β2 recombinant protein exerts the bipolar regulation on the proliferation of the supraneural myeloid body cells (SMB cells) in the quiescent and LPS-activated state, while plays an inhibitory role in the proliferation of quiescent and activated leukocytes in lampreys. Furthermore, caspase-3/7 activity analysis indicates that the lamprey TGF-β2 protects SMB cells from apoptosis after serum deprivation, in contrast to promoting apoptosis of leukocytes. Our composite results offer valuable clues to the origin and evolution of the TGF-β subfamily and imply that TGF-βs are among the most ancestral immune regulators in vertebrates.

Lamprey possess both V1R and V2R olfactory receptors, but only V1Rs are expressed in olfactory sensory neurons

The sense of smell employs some of the largest gene families in the genome to detect and distinguish a multitude of different odors. Within vertebrates, 4 major olfactory receptor families have been described; of which, only 3 (OR, TAAR-like, and V1R) were found already in lamprey, a jawless vertebrate. The forth family (V2R) was believed to have originated later, in jawed vertebrates. Here we have delineated the entire vomeronasal receptor repertoire in 3 lamprey species. We report the presence of 6 v1r and 2 v2r genes in Lethenteron camtschaticum, arctic lamprey, and Lampetra fluviatilis, river lamprey (6 and 1, respectively, in sea lamprey, Petromyzon marinus). Three v1r genes but no v2r genes were found to be expressed in olfactory sensory neurons in the characteristic sparse expression pattern. Our results show the olfactory function of some V1Rs already in lamprey and, unexpectedly, an early origin of the V2R family in the shared ancestor of jawed and jawless vertebrates. However, lamprey v2r genes appear not to have acquired an olfactory function yet, thus dissociating the evolutionary origin of the family from the onset of a function as olfactory receptor.