Tissue-specific antioxidative response and metabolism of paralytic shellfish toxins in scallop (Chlamys farreri) mantle with Alexandrium dinoflagellate exposure.

Bivalves show remarkable capacity to acclimate paralytic shellfish toxins (PSTs) produced by dinoflagellates, severely affecting fishery industry and public health. Here, transcriptomic response to PSTs-producing dinoflagellate (Alexandrium minutum) was investigated in Zhikong scallop (Chlamys farreri) mantle. The PSTs accumulated in C. farreri mantle continually increased during the 15 days exposure, with "oxidation-reduction" genes induced compared to the control group at the 1st and 15th day. Through gene co-expression network analysis, 16 PSTs-responsive modules were enriched with up- or down-regulated genes. The concentration of GTXs, major PSTs in A. minutum and accumulated in scallops, was correlated with the up-regulated magenta module, enriching peroxisome genes as the potential mantle-specific PSTs biomarker. Moreover, Hsp70B2s were inhibited throughout the exposure, which together with the expanded neurotransmitter transporter SLC6As, may play essential roles on neurotransmitter homeostasis in scallop mantle. These results paved the way for a comprehensive understanding of defensive mechanism and homeostatic response in scallop mantle against PSTs.

Transcriptome Analysis Reveals the Genes Involved in Oxidative Stress Responses of Scallop to PST-Producing Algae and a Candidate Biomarker for PST Monitoring.

Paralytic shellfish toxins (PST) could be accumulated in bivalves and cause safety problems. To protect public health, bivalves are examined for PST contamination before entering the market, usually by high-performance liquid chromatography (HPLC) or LC-tandem mass spectrometry (LC-MS/MS) in the lab, which needs PST standards not all available and is time-consuming for large sample sizes. To detect PST toxicity in bivalves rapidly and sensitively, a biomarker gene is highly demanded, but the related study is very limited. In this study, we fed a commercially important bivalve, Patinopecten yessoensis, with the PST-producing dinoflagellate Alexandrium catenella. After 1, 3, and 5 days of exposure, both PST concentrations and toxicity levels in the digestive gland continuously increased. Transcriptome analysis revealed that the differentially expressed genes were significantly enriched in oxidation-reduction process, which included the cytochrome P450 genes (CYPs), type I iodothyronine deiodinase (IOD1s), peroxidasin (PXDN), and acyl-Coenzyme A oxidase 1 (ACOX1) at day 1 and a superoxide dismutase (SOD) at day 5, highlighting the crucial roles of these genes in response to oxidative stress induced by PST. Among the 33 continuously upregulated genes, five showed a significant correlation between gene expression and PST concentration, with the highest correlation present in PyC1QL4-1, the gene encoding Complement C1Q-like protein 4, C1QL4. In addition, the correlation between PyC1QL4-1 expression and PST toxicity was also the highest. Further analysis in another aquaculture scallop (Chlamys farreri) indicated that the expression of CfC1QL4-1, the homolog of PyC1QL4-1, also exhibited significant correlations with both PST toxicity and concentration. Our results reveal the gene expression responses of scallop digestive glands to PST-producing algae and indicate that the C1QL4-1 gene might be a potential biomarker for PST monitoring in scallops, which may provide a convenient way for the early warning and sensitive detection of PST contamination in the bivalves.

Transcriptome and Network Analyses Reveal the Gene Set Involved in PST Accumulation and Responses to Toxic Alexandrium minutum Exposure in the Gills of Chlamys farreri.

Bivalve molluscs are filter-feeding organisms that can accumulate paralytic shellfish toxins (PST) through ingesting toxic marine dinoflagellates. While the effects of PST accumulation upon the physiology of bivalves have been documented, the underlying molecular mechanism remains poorly understood. In this study, transcriptomic analysis was performed in the gills of Zhikong scallop (Chlamys farreri) after 1, 3, 5, 10, and 15 day(s) exposure of PST-producing dinoflagellate Alexandrium minutum. Higher numbers of differentially expressed genes (DEGs) were detected at day 1 (1538) and day 15 (989) than that at day 3 (77), day 5 (82), and day 10 (80) after exposure, and most of the DEGs were only regulated at day 1 or day 15, highlighting different response mechanisms of scallop to PST-producing dinoflagellate at different stages of exposure. Functional enrichment results suggested that PST exposure induced the alterations of nervous system development processes and the activation of xenobiotic metabolism and substance transport processes at the acute and chronic stages of exposure, respectively, while the immune functions were inhibited by PST and might ultimately cause the activation of apoptosis. Furthermore, a weighted gene co-expression network was constructed, and ten responsive modules for toxic algae exposure were identified, among which the yellow module was found to be significantly correlated with PST content. Most of the hub genes in the yellow module were annotated as solute carriers (SLCs) with eight being OCTN1s, implying their dominant roles in regulating PST accumulation in scallop gills. Overall, our results reveal the gene set responding to and involved in PST accumulation in scallop gills, which will deepen our understanding of the molecular mechanism of bivalve resistance to PST.

Identification, characterization and expression analyses of PC4 genes in Yesso scallop (Patinopecten yessoensis) reveal functional differentiations in response to ocean acidification.

Transcriptional coactivator p15 (PC4), considered a multifunctional chromosome associated protein, is actively involved in transcription regulation, DNA replication, damage repair and chromosome formation. Although studies have reported significant effects of PC4 in most vertebrates and some invertebrates, the complete PC4 gene members are less systematically identified and characterized in scallops. In this study, seven PC4 genes (PyPC4s) were identified in the Yesso scallop Patinopecten yessoensis using whole-genome scanning via bioinformatic analyses. Phylogenetic and protein structural analyses were performed to determine the identities and evolutionary relationships of the seven genes. Expression profiles of PyPC4s were further investigated in embryos/larvae at all developmental stages, healthy adult tissues, and mantles that were exposed to low pH stress (pH 6.5 and 7.5) with different time durations (3, 6, 12 and 24 h). Spatiotemporal expression patterns indicated the functional roles of PyPC4s at all development stages and in healthy adult tissues, with PY-3235.33 demonstrating remarkably high constitutive expressions. Expression regulations (up- and down-regulation) of PyPC4s under low pH stress levels demonstrated a time-dependent pattern with functional complementation and/or enhancement, revealing that PyPC4s exhibited differentiated functions in response to ocean acidification (OA). Collectively, our data offer a novel perspective stating that low pH is a potential inducer leading to functional differentiation of PyPC4s in scallops. The results provide preliminary information on the versatile roles of PC4(s) in bivalves in response to OA.

Toxin- and species-dependent regulation of ATP-binding cassette (ABC) transporters in scallops after exposure to paralytic shellfish toxin-producing dinoflagellates.

ATP-binding cassette (ABC) transporters are membrane-bound proteins involved in exporting various xenobiotic compounds from living cells. Bivalve mollusks can accumulate large amounts of paralytic shellfish toxins (PSTs) from marine dinoflagellates. For aquatic invertebrates, the importance of ABC proteins in multi-xenobiotic resistance has been demonstrated, however, the systematic identification of ABC transporters is very limited. In this study, 64 and 67 ABC genes containing all eight described subfamilies (A to H) were identified in Yesso scallop (Patinopecten yessoensis) and Zhikong scallop (Chlamys farreri), respectively, with massive gene expansion being observed in the ABCC and ABCG subfamilies. The kidney harbored more specifically expressed ABC genes than other organs/tissues, most of which belonged to ABCB, ABCC, and ABCG subfamilies. After feeding the scallops with PST-producing dinoflagellates, the expression of scallop ABC genes in the kidney was regulated in toxin- and species-dependent manners. In total, 20 and 24 ABC genes in Zhikong scallop (CfABCs) were induced after exposure to Alexandrium minutum and A. catenella, with the up-regulated members from both ABCC and ABCG subfamilies mainly showing acute and chronic induction by A. minutum and A. catenella, respectively, while the up-regulated CfABCBs mainly showing chronic induction by both dinoflagellates. In Yesso scallop, only eight ABC genes (PyABCs) were regulated after A. catenella exposure, and all the five up-regulated PyABCs were acutely induced. Our findings imply the functional diversity of scallop ABC genes in coping with PST accumulation, which may contribute to the lineage-specific adaptation of scallops for dealing with algal toxins challenge.

Biokinetics and biotransformation of paralytic shellfish toxins in different tissues of Yesso scallops, Patinopecten yessoensis.

Paralytic shellfish toxins (PSTs) are a group of natural toxic substances often found in marine bivalves. Accumulation, anatomical distribution, biotransformation and depuration of PSTs in different tissues of bivalves, however, are still not very well understood. In this study, we investigated biokinetics and biotransformation of PSTs in six different tissues, namely gill, mantle, gonad, adductor muscle, kidney, and digestive gland, in Yesso scallops Patinopecten yessoensis exposed to a toxic strain of dinoflagellate Alexandrium pacificum. High daily accumulation rate (DAR) was recorded at the beginning stage of the experiment. Most of the PSTs in toxic algae ingested by scallops were retained and the toxicity level of PSTs in scallops exceeded the regulatory limit within 5 days. At the late stage of the experiment, however, DAR decreased obviously due to the removal of PSTs. Fitting results of the biokinetics model indicated that the amount of PSTs transferred from digestive gland to mantle, adductor muscle, gonad, kidney, and gill in a decreasing order, and adductor muscle, kidney, and gonad had higher removal rate than gill and mantle. Toxin profile in digestive gland was dominated by N-sulfocarbamoyl toxins 1/2 (C1/2), closely resembled that of the toxic algae. In contrast, toxin components in kidney were dominated by high-potency neosaxitoxin (NEO) and saxitoxin (STX), suggesting that the kidney be a major organ for transformation of PSTs.

Solute carriers in scallop genome: Gene expansion and expression regulation after exposure to toxic dinoflagellate.

The solute carriers (SLCs) are membrane proteins that transport many endogenous and exogenous substances such as xenobiotic toxins. Bivalve mollusks, mainly feeding on microalgae, show marked capacity to accumulate paralytic shellfish toxins (PSTs), the most common and hazardous marine biotoxins produced by dinoflagellates. Exploring the SLCs related to PST accumulation in bivalve could benefit our understanding about the mechanisms of PST bioavailability in bivalve and the adaptations of these species. Herein, we provided the first systematic analysis of SLC genes in mollusks, which identified 673 SLCs (PySLCs, 48 subfamilies) in Yesso scallop (Patinopecten yessoensis), 510 (48 subfamilies) in Pacific oyster (Crassostrea gigas), and 350 (47 subfamilies) in gastropod owl limpet (Lottia gigantea). Significant expansion of subfamilies SLC5, SLC6, SLC16, and SLC23 in scallop, and SLC46 subfamily in both scallop and oyster were revealed. Different PySLC members were highly expressed in the developmental stages and adult tissues, and hepatopancreas harboured more specifically expressed PySLCs than other tissues/organs. After feeding the scallops with PST-producing dinoflagellate, 131 PySLCs were regulated and more than half of them were from the expanded subfamilies. The trend of expression fold change in regulated PySLCs was consistent with that of PST changes in hepatopancreas, implying the possible involvement of these PySLCs in PST transport and homeostasis. In addition, the PySLCs from the expanded subfamily were revealed to be under positive selection, which might be related to lineage-specific adaptation to the marine environments with algae derived biotoxins.

Tissue-Biased and Species-Specific Regulation of Glutathione Peroxidase (GPx) Genes in Scallops Exposed to Toxic Dinoflagellates.

Marine bivalves could accumulate paralytic shellfish toxins (PSTs) produced by toxic microalgae, which might induce oxidative stress. Glutathione peroxidases (GPxs) are key enzymes functioning in the antioxidant defense, whereas our understanding of their roles in PST challenge in bivalves is limited. Herein, through genome-wide screening, we identified nine (CfGPx) and eight (PyGPx) GPx genes in Zhikong scallop (Chlamys farreri) and Yesso scallop (Patinopecten yessoensis), respectively, and revealed the expansion of GPx3 sub-family in both species. RNA-Seq analysis revealed high expression of scallop GPx3s after D stage larva during early development, and in adult hepatopancreas. However, in scallops exposed to PST-producing dinoflagellates, no GPx was significantly induced in the hepatopancreas. In scallop kidneys where PSTs were transformed to higher toxic analogs, most CfGPxs were up-regulated, with CfGPx3s being acutely and chronically induced by Alexandrium minutum and A. catenella exposure, respectively, but only one PyGPx from GPx3 subfamily was up-regulated by A. catenella exposure. Our results suggest the function of scallop GPxs in protecting kidneys against the oxidative stresses by PST accumulation or transformation. The tissue-, species-, and toxin-dependent expression pattern of scallop GPxs also implied their functional diversity in response to toxin exposure.

Genome-Wide Identification and Characterization of SODs in Zhikong Scallop Reveals Gene Expansion and Regulation Divergence after Toxic Dinoflagellate Exposure.

As filter-feeding animals mainly ingesting microalgae, bivalves could accumulate paralytic shellfish toxins (PSTs) produced by harmful algae through diet. To protect themselves from the toxic effects of PSTs, especially the concomitant oxidative damage, the production of superoxide dismutase (SOD), which is the only eukaryotic metalloenzyme capable of detoxifying superoxide, may assist with toxin tolerance in bivalves. To better understand this process, in the present study, we performed the first systematic analysis of SOD genes in bivalve Chlamys farreri, an important aquaculture species in China. A total of six Cu/Zn-SODs (SOD1-6) and two Mn-SODs (SOD7, SOD8) were identified in C. farreri, with gene expansion being revealed in Cu/Zn-SODs. In scallops exposed to two different PSTs-producing dinoflagellates, Alexandrium minutum and A. catenella, expression regulation of SOD genes was analyzed in the top ranked toxin-rich organs, the hepatopancreas and the kidney. In hepatopancreas, which mainly accumulates the incoming PSTs, all of the six Cu/Zn-SODs showed significant alterations after A. minutum exposure, with SOD1, 2, 3, 5, and 6 being up-regulated, and SOD4 being down-regulated, while no significant change was detected in Mn-SODs. After A. catenella exposure, up-regulation was observed in SOD2, 4, 6, and 8, and SOD7 was down-regulated. In the kidney, where PSTs transformation occurs, SOD4, 5, 6, and 8 were up-regulated, and SOD7 was down-regulated in response to A. minutum feeding. After A. catenella exposure, all the Cu/Zn-SODs except SOD1 were up-regulated, and SOD7 was down-regulated in kidney. Overall, in scallops after ingesting different toxic algae, SOD up-regulation mainly occurred in the expanded Cu/Zn-SOD group, and SOD6 was the only member being up-regulated in both toxic organs, which also showed the highest fold change among all the SODs, implying the importance of SOD6 in protecting scallops from the stress of PSTs. Our results suggest the diverse function of scallop SODs in response to the PST-producing algae challenge, and the expansion of Cu/Zn-SODs might be implicated in the adaptive evolution of scallops or bivalves with respect to antioxidant defense against the ingested toxic algae.

Alternative splicing, spatiotemporal expression of TEP family genes in Yesso scallop (Patinopecten yessoensis) and their disparity in responses to ocean acidification.

The complement system constitutes a highly sophisticated and powerful body defense machinery acting in the innate immunity of both vertebrates and invertebrates. As central components of the complement system, significant effects of thioester-containing protein (TEP) family members on immunity have been reported in most vertebrates and in some invertebrates, but the spatiotemporal expression and regulatory patterns of TEP family genes under environmental stress have been less widely investigated in scallops. In this study, expression profiling of TEP family members in the Yesso scallop Patinopecten yessoensis (designated PyTEPs) was performed at all developmental stages, in different healthy adult tissues, and in mantles during exposure to different levels of acidification (pH = 6.5 and 7.5) for different time points (3, 6, 12 and 24 h); this profiling was accomplished through in silico analysis of transcriptome and genome databases. Spatiotemporal expression patterns revealed that PyTEPs had specific functional differentiation in all stages of growth and development of the scallop. Expression analysis confirmed the inducible expression patterns of PyTEPs during exposure to acidification. Gene duplication and alternative splicing events simultaneously occurred in PyTEP1. Seven different cDNA variants of PyTEP1 (designated PyTEP1-A-PyTEP1-G) were identified in the scallop mantle transcriptome during acidic stress. These variants were produced by the alternative splicing of seven differentially transcribed exons (exons 18-24), which encode the highly variable central region. The responses to immune stress may have arisen through the gene duplication and alternative splicing of PyTEP1. The sequence diversity of PyTEP1 isoforms and their different expression profiles in response to ocean acidification (OA) suggested a mechanism used by scallops to differentiate and regulate PyTEP1 gene expression. Collectively, these results demonstrate the gene duplication and alternative splicing of TEP family genes and provide valuable resources for elucidating their versatile roles in bivalve innate immune responses to OA challenge.

Diverse expression regulation of Hsp70 genes in scallops after exposure to toxic Alexandrium dinoflagellates.

Heat shock proteins 70KD (Hsp70s) are highly conserved molecular chaperones with essential roles against biotic and abiotic stressors. Marine bivalves inhabit highly complex environments and could accumulate paralytic shellfish toxins (PSTs), the well-noted neurotoxins generated during harmful algal blooms. Here, we systematically analyzed Hsp70 genes (CfHsp70s) in Zhikong scallop (Chlamys farreri), an important aquaculture mollusk in China. Sixty-five CfHsp70s from eight sub-families were identified, and 47 of these genes showed expansion in the Hspa12 sub-family. After exposure to different PST-producing dinoflagellates, Alexandrium minutum and Alexandrium catenella, diverse CfHsp70s regulation presented in scallop hepatopancreas, mainly accumulating incoming PSTs, and kidneys, transforming PSTs into higher toxic analogs. All the up-regulated CfHsp70s were from CfHsp70B2, CfHspa12, and CfHspa5 sub-families. CfHsp70B2 sub-family was mainly induced in the hepatopancreas, and CfHspa12 sub-family was highly induced in the kidneys. CfHsp70s up-regulation under two dinoflagellates exposure was stronger in the kidneys (log2FC: 19.5 and 18.6) than that in hepatopancreas (log2FC: 4.3 and 6.1). Exposure to different Alexandrium species had varying effects, that in hepatopancreas, CfHsp70B2s were chronically induced only after A. catenella exposure, whereas in kidney, CfHspa12s were more acutely induced after exposure of A. minutum than A. caenella. Moreover, in Yesso scallops (Patinopecten yessoensis), only Hspa12s were up-regulated in hepatopancreas after A. catenella exposure, and all the Hsp70B2s were down-regulated. These organ-, toxin-, and species-dependent Hsp70 regulation suggested the functional diversity of duplicated Hsp70s in response to the stress by PST-producing algae. Our findings provide insights into the evolution and functional characteristics of Hsp70s in scallops.

A carotenoid oxygenase is responsible for muscle coloration in scallop.

As lipid microconstituents mainly of plant origin, carotenoids are essential nutrients for humans and animals, and carotenoid coloration represents an important meat quality parameter for many farmed animals. Currently, the mechanism of carotenoid bioavailability in animals is largely unknown mainly due to the limited approaches applied, the shortage of suitable model systems and the restricted taxonomic focus. The mollusk Yesso scallop (Patinopecten yessoensis) possessing orange adductor muscle with carotenoid deposition, provides a unique opportunity to research the mechanism underlying carotenoid utilization in animals. Herein, through family construction and analysis, we found that carotenoid coloration in scallop muscle is inherited as a recessive Mendelian trait. Using a combination of genomic approaches, we mapped this trait onto chromosome 8, where PyBCO-like 1 encoding carotenoid oxygenase was the only differentially expressed gene between the white and orange muscles (FDR = 2.75E-21), with 11.28-fold downregulation in the orange muscle. Further functional assays showed that PyBCO-like 1 is capable of degrading ß-carotene, and inhibiting PyBCO-like 1 expression in the white muscle resulted in muscle coloration and carotenoid deposition. In the hepatopancreas, which is the organ for digestion and absorption, neither the scallop carotenoid concentration nor PyBCO-like 1 expression were significantly different between the two scallops. These results indicate that carotenoids could be taken up in both white- and orange-muscle scallops and then degraded by PyBCO-like 1 in the white muscle. Our data suggest that PyBCO-like 1 is the essential gene for carotenoid metabolism in scallop muscle, and its downregulation leads to carotenoid deposition and muscle coloration.

Sea cucumber genome provides insights into saponin biosynthesis and aestivation regulation.

Echinoderms exhibit several fascinating evolutionary innovations that are rarely seen in the animal kingdom, but how these animals attained such features is not well understood. Here we report the sequencing and analysis of the genome and extensive transcriptomes of the sea cucumber Apostichopus japonicus, a species from a special echinoderm group with extraordinary potential for saponin synthesis, aestivation and organ regeneration. The sea cucumber does not possess a reorganized Hox cluster as previously assumed for all echinoderms, and the spatial expression of Hox7 and Hox11/13b potentially guides the embryo-to-larva axial transformation. Contrary to the typical production of lanosterol in animal cholesterol synthesis, the oxidosqualene cyclase of sea cucumber produces parkeol for saponin synthesis and has "plant-like" motifs suggestive of convergent evolution. The transcriptional factors Klf2 and Egr1 are identified as key regulators of aestivation, probably exerting their effects through a clock gene-controlled process. Intestinal hypometabolism during aestivation is driven by the DNA hypermethylation of various metabolic gene pathways, whereas the transcriptional network of intestine regeneration involves diverse signaling pathways, including Wnt, Hippo and FGF. Decoding the sea cucumber genome provides a new avenue for an in-depth understanding of the extraordinary features of sea cucumbers and other echinoderms.

Identification and characterization of TEP family genes in Yesso scallop (Patinopecten yessoensis) and their diverse expression patterns in response to bacterial infection.

Thioester-containing protein (TEP) family members are characterized by their unique intrachain ß-cysteinyl-γ-glutamyl thioesters, and they play important roles in innate immune responses. Although significant effects of TEP members on immunity have been reported in most vertebrates, as well as certain invertebrates, the complete TEP family has not been systematically characterized in scallops. In this study, five TEP family genes (PyC3, PyA2M, PyTEP1, PyTEP2 and PyCD109) were identified from Yesso scallop (Patinopecten yessoensis) through whole-genome scanning, including one pair of tandem duplications located on the same scaffold. Phylogenetic and protein structural analyses were performed to determine the identities and evolutionary relationships of the five genes (PyTEPs). The vast distribution of PyTEPs in TEP subfamilies confirmed that the Yesso scallop contains relatively comprehensive types of TEP members in evolution. The expression profiles of PyTEPs were determined in hemocytes after bacterial infection with gram-positive (Micrococcus luteus) and gram-negative (Vibrio anguillarum) using quantitative real-time PCR (qRT-PCR). Expression analysis revealed that the PyTEP genes exhibited disparate expression patterns in response to the infection by gram bacteria. A majority of PyTEP genes were overexpressed after bacterial stimulation at most time points, especially the notable elevation displayed by duplicated genes after V. anguillarum challenge. Interestingly, at different infection times, PyTEP1 and PyTEP2 shared analogous expression patterns, as did PyC3 and PyCD109. Taken together, these results help to characterize gene duplication and the evolutionary origin of PyTEPs and supplied valuable resources for elucidating their versatile roles in bivalve innate immune responses to bacterial pathogen challenges.

Scallop genome reveals molecular adaptations to semi-sessile life and neurotoxins.

Bivalve molluscs are descendants of an early-Cambrian lineage superbly adapted to benthic filter feeding. Adaptations in form and behavior are well recognized, but the underlying molecular mechanisms are largely unknown. Here, we investigate the genome, various transcriptomes, and proteomes of the scallop Chlamys farreri, a semi-sessile bivalve with well-developed adductor muscle, sophisticated eyes, and remarkable neurotoxin resistance. The scallop's large striated muscle is energy-dynamic but not fully differentiated from smooth muscle. Its eyes are supported by highly diverse, intronless opsins expanded by retroposition for broadened spectral sensitivity. Rapid byssal secretion is enabled by a specialized foot and multiple proteins including expanded tyrosinases. The scallop uses hepatopancreas to accumulate neurotoxins and kidney to transform to high-toxicity forms through expanded sulfotransferases, probably as deterrence against predation, while it achieves neurotoxin resistance through point mutations in sodium channels. These findings suggest that expansion and mutation of those genes may have profound effects on scallop's phenotype and adaptation.

Scallop genome provides insights into evolution of bilaterian karyotype and development.

Reconstructing the genomes of bilaterian ancestors is central to our understanding of animal evolution, where knowledge from ancient and/or slow-evolving bilaterian lineages is critical. Here we report a high-quality, chromosome-anchored reference genome for the scallop Patinopecten yessoensis, a bivalve mollusc that has a slow-evolving genome with many ancestral features. Chromosome-based macrosynteny analysis reveals a striking correspondence between the 19 scallop chromosomes and the 17 presumed ancestral bilaterian linkage groups at a level of conservation previously unseen, suggesting that the scallop may have a karyotype close to that of the bilaterian ancestor. Scallop Hox gene expression follows a new mode of subcluster temporal co-linearity that is possibly ancestral and may provide great potential in supporting diverse bilaterian body plans. Transcriptome analysis of scallop mantle eyes finds unexpected diversity in phototransduction cascades and a potentially ancient Pax2/5/8-dependent pathway for noncephalic eyes. The outstanding preservation of ancestral karyotype and developmental control makes the scallop genome a valuable resource for understanding early bilaterian evolution and biology.

Genome-wide identification, characterization and expression analyses of TLRs in Yesso scallop (Patinopecten yessoensis) provide insight into the disparity of responses to acidifying exposure in bivalves.

Toll-like receptors (TLRs) play a crucial role in innate immunity by recognizing specific pathogen-associated molecular patterns, including lipoproteins, lipopeptides, lipopolysaccharide, flagellin, dsRNA, ssRNA and CpG DNA motifs. Although significant effects of TLRs on immunity have been reported in most vertebrates and some invertebrates, the complete TLR superfamily has not been systematically characterized in scallops. In this study, 18 TLR genes were identified from Yesso scallop (Patinopecten yessoensis) using whole-genome scanning. Phylogenetic and protein structural analyses were performed to determine the identities and evolutionary relationships of the 18 genes. Extensive expansion of TLR genes from the Yesso scallop genome indicated gene duplication events. In addition, expression profiling of PyTLRs was performed at different acidifying exposure levels (pH = 6.50, 7.50) with different challenge durations (3, 6, 12 and 24 h) via in silico analysis using transcriptome and genome databases. Our results confirmed the inducible expression patterns of PyTLRs under acidifying exposure, and the responses to immune stress may have arisen through adaptive recruitment of tandem duplications of TLR genes. Collectively, this study provides novel insight into PyTLRs as well as the specific role and response of TLR signaling pathways in host immune responses against acidifying exposure in bivalves.

Hsp70 gene expansions in the scallop Patinopecten yessoensis and their expression regulation after exposure to the toxic dinoflagellate Alexandrium catenella.

Heat shock protein 70 (Hsp70s) family members are present in virtually all living organisms and perform a fundamental role against different types of environmental stressors and pathogenic organisms. Marine bivalves live in highly dynamic environments and may accumulate paralytic shellfish toxins (PSTs), a class of well-known neurotoxins closely associated with harmful algal blooms (HABs). Here, we provide a systematic analysis of Hsp70 genes (PyHsp70s) in the genome of Yesso scallop (Patinopecten yessoensis), an important aquaculture species in China, through in silico analysis using transcriptome and genome databases. Phylogenetic analyses indicated extensive expansion of Hsp70 genes from the Hspa12 sub-family in the Yesso scallop and also the bivalve lineages, with gene duplication events before or after the split between the Yesso scallop and the Pacific oyster. In addition, we determined the expression patterns of PyHsp70s after exposure to Alexandrium catenella, the dinoflagellate producing PSTs. Our results confirmed the inducible expression patterns of PyHsp70s under PSTs stress, and the responses to the toxic stress may have arisen through the adaptive recruitment of tandem duplication of Hsp70 genes. These findings provide a thorough overview of the evolution and modification of the Hsp70 family, which will gain insights into the functional characteristics of scallop Hsp70 genes in response to different stresses.

The Rho GTPase Family Genes in Bivalvia Genomes: Sequence, Evolution and Expression Analysis.

BACKGROUND: Rho GTPases are important members of the Ras superfamily, which represents the largest signaling protein family in eukaryotes, and function as key molecular switches in converting and amplifying external signals into cellular responses. Although numerous analyses of Rho family genes have been reported, including their functions and evolution, a systematic analysis of this family has not been performed in Mollusca or in Bivalvia, one of the most important classes of Mollusca. RESULTS: In this study, we systematically identified and characterized a total set (Rho, Rac, Mig, Cdc42, Tc10, Rnd, RhoU, RhoBTB and Miro) of thirty Rho GTPase genes in three bivalve species, including nine in the Yesso scallop Patinopecten yessoensis, nine in the Zhikong scallop Chlamys farreri, and twelve in the Pacific oyster Crassostrea gigas. Phylogenetic analysis and interspecies comparison indicated that bivalves might possess the most complete types of Rho genes in invertebrates. A multiple RNA-seq dataset was used to investigate the expression profiles of bivalve Rho genes, revealing that the examined scallops share more similar Rho expression patterns than the oyster, whereas more Rho mRNAs are expressed in C. farreri and C. gigas than in P. yessoensis. Additionally, Rho, Rac and Cdc42 were found to be duplicated in the oyster but not in the scallops. Among the expanded Rho genes of C. gigas, duplication pairs with high synonymous substitution rates (Ks) displayed greater differences in expression. CONCLUSION: A comprehensive analysis of bivalve Rho GTPase family genes was performed in scallop and oyster species, and Rho genes in bivalves exhibit greater conservation than those in any other invertebrate. This is the first study focusing on a genome-wide characterization of Rho GTPase genes in bivalves, and the findings will provide a valuable resource for a better understanding of Rho evolution and Rho GTPase function in Bivalvia.