Water temperature effects on embryonic development and GnRH-like expression in kisslip cuttlefish (Sepia lycidas) hinting the potential role of GnRH-like peptide.

Gonadotropin-releasing hormone (GnRH)-like peptides are multifunctional neuropeptides involved in cardiac control, early ontogenesis, and reproduction in cephalopods. However, the precise role of GnRH-like peptides in embryonic development and juvenile growth in cephalopods remains unknown. In this study, we showed that GnRH-like peptides are involved in the embryonic development of kisslip cuttlefish (Sepia lycidas). We confirmed that higher water temperatures induced early hatching. Simultaneously, we found that brain GnRH-like peptide gene expression gradually increased with increasing hatching speed. However, the rise in water temperature within a suitable range had no effect on the juvenile sex ratio or early gonadal development. Our results indicate that GnRH-like peptides may play an accelerating role in embryonic development; however, they are not involved in sex determination or early gonadal development in kisslip cuttlefish.

Expression profile of GnRH-like peptide during gonadal sex differentiation in the cephalopod kisslip cuttlefish, Sepia lycidas.

Gonadotropin-releasing hormone (GnRH) is one of the most important neuroendocrine regulators for animal reproduction. GnRH-like peptide (GnRH-like) has recently been shown to play a critical reproductive role mainly in gametogenesis or steroidogenesis in the gonads of some molluscs, including cephalopods. However, its involvement in gonadal sex differentiation remains unknown. Here, we show the expression profile of GnRH-like in the brain of the cephalopod kisslip cuttlefish, Sepia lycidas, throughout gonadal sex differentiation, by quantitative real time RT-PCR and immunohistochemistry. We found that GnRH-like could be detected in the brain at a sexually undifferentiated stage, and its expression level significantly increased upon initiation of gonadal sex differentiation. However, no significant difference in GnRH-like expression levels was observed between sexes during gonadal sex differentiation. Additionally, we demonstrated immunoreactivity of GnRH-like in glial cells or immature neurons, which are mainly distributed in the non-reproductive related area of the cephalopod brain, suggesting the immature function of the reproductive endocrine axis during early ontogenesis. Our results demonstrate for the first time, the expression profile of GnRH-like during early ontogenesis in cephalopods.

Hepatic expression profiles of three subtypes of vitellogenin and estrogen receptor during vitellogenesis in cultured female yellowtail.

Estradiol-17ß (E2) promotes the transcription of vitellogenin (Vtg) via nuclear estrogen receptor (ER). Three Vtg (VtgAa, VtgAb, and VtgC) and ER subtypes (ERα, ERß1, and ERß2) have been reported in perciform fish; however, the relationship between the transcriptional regulation of Vtg and ER subtypes remains unclear. Molecular characterization was performed and the expression profiles of vtg and er subtypes were investigated to elucidate mechanisms of synthesis of vtg subtypes in yellowtail, Seriola quinqueradiata. Primary structures and promoter regions were revealed in three subtypes of vtg and er, and all the vtg subtypes and erα were presumed to be estrogen-responsive genes. When all vtg subtypes were expressed significantly in the liver, hepatic expression levels of all the er subtypes also increased. Conversely, although plasma E2 concentrations did not change significantly, the concentrations were high at the same time. Hepatic expression levels of all the vtg subtypes were highly correlated with hepatic erα, rather than with hepatic erß subtypes and plasma E2. A high positive correlation was also observed between erß1 and ß2, which seemed to be highly expressed at the pre- and late-vitellogenic stages. The results of the present study suggest that the transcription of the three vtg subtypes are regulated by three ER subtypes jointly, and ERα is the key transcription factor regulating the three vtg subtypes in yellowtail.

Development of a giant grouper Luteinizing Hormone (LH) Enzyme-Linked Immunosorbent Assay (ELISA) and its use towards understanding sexual development in grouper.

A recombinant giant grouper Luteinizing Hormone (LH) consisting of tethered beta and alpha subunits was produced in a yeast expression system. The giant grouper LH ß-subunit was also produced and administered to rabbits for antibody development. The recombinant LH and its antibody were used to develop an Enzyme Linked Immunosorbent Assay (ELISA). This ELISA enabled detection of plasma LH levels in groupers at a sensitivity between 391 pg/ml and 200 ng/ml. Different species of grouper were assayed with this ELISA in conjunction with gonadal histology and body condition data to identify links between circulating LH levels and sexual development. We found that circulating levels of LH decreased when oocytes began to degenerate, and sex-transition gonadal characteristics were apparent when LH levels decreased further. When circulating LH levels were related to body condition (body weight/ body length), transitioning-stage fish had relatively high body condition but low plasma LH levels. This observation was similar across multiple grouper species and indicates that plasma LH levels combined with body condition may be a marker for early male identification in the protogynous hermaphrodite groupers.

Development of specific chemiluminescent immunoassays for three subtypes of vitellogenin in grey mullet (Mugil cephalus).

Chemiluminescent immunoassays (CLIAs) were developed for each of three subtypes of vitellogenin (VtgAa, VtgAb and VtgC) in grey mullet, primarily for use in monitoring estrogenic pollution of the environment. The working range of VtgAa-CLIA and VtgAb-CLIA was from 0.975 to 1,000 ng/ml, while that of VtgC-CLIA was from 0.487 to 1,000 ng/ml. Each CLIA appeared to be specific to the targeted Vtg subtype. Intra- and inter-assay coefficients of variation in the developed CLIAs were lower than 10%. In male serum, VtgAa, VtgAb and VtgC were detected in ranges from 0.01 to 0.38, 0.02 to 1.01, and 0.01 to 3.12 µg/ml, respectively, during various sampling periods. In vitellogenic females (October), serum VtgAb levels (1,192.05 ±â€¯237.81 µg/ml) were significantly higher than levels of the other two Vtg subtypes (120.82 ±â€¯30.42 and 119.23 ±â€¯16.95 µg/ml for VtgAa and VtgC, respectively). When immature mullet were fed diets containing 17α-ethinylestradiol (EE2) at three different doses (0.4, 40 and 4,000 ng/g body weight), all Vtg subtypes were induced by 40 ng/g and 4,000 ng/g EE2. The VtgC (610.30 ±â€¯150.18 µg/ml) was most highly expressed among the three Vtgs in fish fed 40 ng/g EE2, while VtgAb (33.25 ±â€¯13.58 mg/ml) was highest in expression in fish fed 4,000 ng/g EE2. The present study provided practical subtype-specific Vtg assays for the first time in grey mullet, providing the necessary means to evaluate estrogenic activities in aquatic environments.

Photoperiodic regulation of plasma gonadotropin levels in previtellogenic greater amberjack (Seriola dumerili).

In Seriola species, exposure to a long photoperiod regime is known to induce ovarian development. This study examined photoperiodic effects on pituitary gene expression and plasma levels of follicle-stimulating hormone (Fsh) and luteinizing hormone (Lh) in previtellogenic greater amberjack (Seriola dumerili). The fish were exposed to short (8L:16D) or long (18L:6D) photoperiod. The water temperature was maintained at 22 °C. Compared with the short-photoperiod group, plasma Fsh levels were higher on days 10 and 30 in the long-photoperiod group, but plasma Lh levels did not significantly differ. On day 30, pituitary Fsh- and Lh-ß subunit gene expressions were also higher in the long-photoperiod group than the short-photoperiod group, whereas α-subunit gene expressions were higher on days 20 and 30. Throughout the experiment, average gonadosomatic index and plasma E2 levels did not significantly differ between the two groups. This study clearly demonstrated that a long photoperiod induced Fsh release in the previtellogenic fish followed by upregulation of pituitary Fsh and Lh subunit gene expressions. An increase in plasma Fsh levels may be a key factor that mediates the photoperiodic effect on the initiation of ovarian development.

Standardization of plasma vitellogenin level for surveying environmental estrogen pollution using the Japanese common goby Acanthogobius flavimanus.

Field surveys of the impact of environmental estrogen (EE) pollution in aquatic wildlife have been conducted using vitellogenin (VTG) as a biomarker to evaluate the influence of EE. However, a standard baseline of VTG level that can be used to evaluate EE pollution has not been fully determined. To the best of our knowledge, the present study is the first to determine the standard baseline VTG level for evaluating the biological effects of EE pollution using the Japanese common goby (Acanthogobius flavimanus) as the target model fish. Plasma VTG and estradiol-17ß (E2 ) levels associated with the reproductive cycle of wild goby inhabiting an unpolluted environment were measured. Mean plasma VTG and E2 levels exhibited similar changes, increasing in the yolk vesicle stage and peaking in the tertiary yolk stage in females. However, plasma VTG and E2 levels showed no significant changes in males, remaining at low levels throughout the reproductive cycle. The highest VTG levels in females and males were 1.6 mg ml-1 and 124.87 ng ml-1 , respectively. These results indicate that the baseline level (normal level) in males was approximately 130 ng ml-1 at most. We concluded that the threshold between normal and abnormal levels with a 10% risk rate was 150 ng ml-1 in the wild male goby. Plasma VTG levels in males captured from Nagasaki Harbor were higher than the threshold in each reproductive developmental stage, indicating the possibility of EE pollution at this site. The biological standard baseline for VTG established in this study is useful for assessing EE pollution in natural waters.

The Mechanism of Low-Temperature Tolerance in Fish.

In this chapter, we cover the life history of fish in low-temperature environments, including their overwintering behavior and the physiological mechanisms by which they maintain life in cold environments, based on research to date. There is relatively little research on low-temperature tolerance of fish, compared with research on this phenomenon in mammals and birds, which are also vertebrates, and the mechanisms in fish have not been fully elucidated. First, we cover the life history of fish that overwinter by entering dormancy or hibernation. Next, we describe the mechanism that controls body temperature in fish that survive low-temperature environments. Finally, we introduce the physiological mechanisms for survival in extremely low-temperature environments, particularly antifreeze proteins.

Changes in gene expression and cellular localization of insulin-like growth factors 1 and 2 in the ovaries during ovary development of the yellowtail, Seriola quinqueradiata.

A method of controlling the somatic growth and reproduction of yellowtail fish (Seriola quinqueradiata) is needed in order to establish methods for the efficient aquaculture production of the species. However, little information about the hormonal interactions between somatic growth and reproduction is available for marine teleosts. There is accumulating evidence that insulin-like growth factor (IGF), a major hormone related somatic growth, plays an important role in fish reproduction. As the first step toward understanding the physiological role of IGF in the development of yellowtail ovaries, we characterized the expression and cellular localization of IGF-1 and IGF-2 in the ovary during development. We histologically classified the maturity of two-year-old females with ovaries at various developmental stages into the perinucleolar (Pn), yolk vesicle (Yv), primary yolk (Py), secondary yolk and tertiary yolk (Ty) stages, according to the most advanced type of oocyte present. The IGF-1 gene expression showed constitutively high levels at the different developmental stages, although IGF-1 mRNA levels tended to increase from the Py to the Ty stage with vitellogenesis, reaching maximum levels during the Ty stage. The IGF-2 mRNA levels increased as ovarian development advanced. Using immunohistochemistry methods, immunoreactive IGF-1 was mainly detected in the theca cells of ovarian follicles during late secondary oocyte growth, and in part of the granulosa cells of Ty stage oocytes. IGF-2 immunoreactivity was observed in all granulosa cells in layer in Ty stage oocytes. These results indicate that follicular IGFs may be involved in yellowtail reproduction via autocrine/paracrine mechanisms.

Greater amberjack Fsh, Lh, and their receptors: Plasma and mRNA profiles during ovarian development.

To understand the endocrine regulation of ovarian development in a multiple spawning fish, the relationship between gonadotropins (Gths; follicle-stimulating hormone [Fsh] and luteinizing hormone [Lh]) and their receptors (Gthrs; Fshr and Lhr) were investigated in greater amberjack (Seriola dumerili). cDNAs encoding the Gth subunits (Fshß, Lhß, and glycoprotein α [Gpα]) and Gthrs were cloned. The in vitro reporter gene assay using recombinant hormones revealed that greater amberjack Fshr and Lhr responded strongly to their own ligands. Competitive enzyme-linked immunosorbent assays (ELISAs) were developed for measuring greater amberjack Fsh and Lh. Anti-Fsh and anti-Lh antibodies were raised against recombinant chimeric single-chain Gths consisting of greater amberjack Fshß (or Lhß) with rabbit GPα. The validation study showed that the ELISAs were precise (intra- and inter-assay coefficient of variation, <10%) and sensitive (detection limit of 0.2ng/ml for Fsh and 0.8ng/ml for Lh) with low cross-reactivity. A good parallelism between the standard curve and serial dilutions of greater amberjack plasma and pituitary extract were obtained. In female greater amberjack, pituitary fshb, ovarian fshr, and plasma E2 gradually increased during ovarian development, and plasma Fsh significantly increased during the post-spawning period. This suggests that Fsh plays a role throughout ovarian development and during the post-spawning period. Pituitary lhb, ovarian lhr, and plasma Lh were high during the spawning period, suggesting that the synthesis and secretion of Lh, and Lhr expression are upregulated to induce final oocyte maturation and ovulation.

Use of a gyroscope/accelerometer data logger to identify alternative feeding behaviours in fish.

We examined whether we could identify the feeding behaviours of the trophic generalist fish Epinephelus ongus on different prey types (crabs and fish) using a data logger that incorporated a three-axis gyroscope and a three-axis accelerometer. Feeding behaviours and other burst behaviours, including escape responses, intraspecific interactions and routine movements, were recorded from six E. ongus individuals using data loggers sampling at 200 Hz, and were validated by simultaneously recorded video images. For each data-logger record, we extracted 5 s of data when any of the three-axis accelerations exceeded absolute 2.0 g, to capture all feeding behaviours and other burst behaviours. Each feeding behaviour was then identified using a combination of parameters that were derived from the extracted data. Using decision trees with the parameters, high true identification rates (87.5% for both feeding behaviours) with low false identification rates (5% for crab-eating and 6.3% for fish-eating) were achieved for both feeding behaviours.

Molecular cloning and characterization of the expression profiles of vitellogenin transcripts in the dojo loach (Misgurnus anguillicaudatus) in response to 17α-ethinylestradiol and 17ß-estradiol administration.

The gene, vitellogenin (vtg) was cloned and characterized in the dojo loach (Misgurnus anguillicaudatus), an indigenous freshwater species in East Asia, in order to develop tools for detecting the effects of estrogenic endocrine-disrupting chemicals (EEDCs). Full-length cDNAs encoding seven distinct vtg transcripts (vtg1-7) were obtained. The corresponding deduced amino acid sequences (Vtg1-7) were divided into two types; type I (Vtg1-6; 89-99% identical), which contained both lipovitellin (Lv) and phosvitin (Pv), and type II (Vtg7), which contained Lv alone. Phylogenetic analysis revealed that the type I and type II Vtgs in the loach could be classified as VtgAo1 and VtgC types, respectively. Immuno-biochemical analyses using type-specific Vtg antisera revealed that VtgAo1 proteins appeared to be the major Vtg type in this species. Males were administered (aqueous exposure) either 17ß-estradiol (E2) or 17α-ethinylestradiol (EE2), the results from which were used to determine that hepatic vtgAo1 expression was estrogen-sensitive. The precise classification of the loach vtg/Vtg products, as well as their induction profiles following the estrogenic stimulation, provide a basis for their use as sensitive biomarkers when EEDC activities are evaluated in the freshwater environments in East Asia.

High water temperature triggers early sexual maturation in the juvenile red spotted grouper Epinephelus akaara: Via regulation of reproduction-related hormones in the brain-pituitary-gonadal axis.

The red spotted grouper Epinephelus akaara is a marine species of economic importance and also at risk of extinction. This study investigated the effects of high water temperature on the growth and maturation of juvenile E. akaara females. From 160-420 days post-hatching (dph), the fish were maintained under natural water temperature (NT) and a constant high-water temperature (HT). From 240 dph, both the total length and body weight in the HT group were greater than in NT group. After 360 dph, the gonadosomatic index was also increased in the HT group compared to NT group. Mature oocytes were only observed in the HT group at 330, 360, and 390 dph. Both kiss1 and kiss2 levels increased at 240 and 270 dph in both groups; however, they were greater in the HT group at 240 dph. Similarly, gpr54 levels after 360 dph were greater in the HT group, suggesting that kisspeptin is related to maturation via its receptor gpr54. Levels of fshß and lhß were greater in the HT group after 360 dph. Estradiol-17ß (E2) levels after 160 dph (except 300 dph) were greater in the HT group than in the NT group, suggesting that the higher E2 levels trigger maturation, and is related to increased fshß and lhß. This study provides evidence that high water temperature is effective in accelerating growth and triggering early maturation of juvenile E. akaara, via regulating gpr54, fshß, lhß, and E2 levels.

Bifenthrin induces changes in clinical poisoning symptoms, oxidative stress, DNA damage, histological characteristics, and transcriptome in Chinese giant salamander (Andrias davidianus) larvae.

Bifenthrin (BF) is a broad-spectrum insecticide that has gained widespread use due to its high effectiveness. However, there is limited research on the potential toxic effects of bifenthrin pollution on amphibians. This study aimed to investigate the 50 % lethal concentration (LC50) and safety concentration of Chinese giant salamanders (CGS) exposed to BF (at 0, 6.25,12.5,25 and 50 µg/L BF) for 96 h. Subsequently, CGS were exposed to BF (at 0, 0.04, and 4 µg/L BF) for one week to investigate its toxic effects. Clinical poisoning symptoms, liver pathology, oxidative stress factors, DNA damage, and transcriptome differences were observed and analyzed. The results indicate that exposure to BF at 4 µg/L significantly decreased the adenosine-triphosphate (ATP), superoxide dismutase (SOD), glutathione (GSH), and catalase (CAT) contents in the brain, liver, and kidney of CGS. Additionally, the study found that the malondialdehyde (MDA), reactive oxygen species (ROS), and 8-hydroxydeoxyguanosine (8-OHdG) contents were increased. The liver tissue exhibited significant inflammatory reactions and structural malformations. RNA-seq analysis of the liver showed that BF caused abnormal antioxidant indices of CGS. This affected molecular function genes such as catalytic activity, ATP-dependent activity, metabolic processes, signaling and immune system processes, behavior, and detoxification, which were significantly upregulated, resulting in the differential genes significantly enriched in the calcium signaling pathway, PPARα signaling pathway and NF-kB signaling pathway. The results suggest that BF induces the abnormal production of free radicals, which overwhelms the body's self-defense system, leading to varying degrees of oxidative stress. This can result in oxidative damage, DNA damage, abnormal lipid metabolism, autoimmune diseases, clinical poisoning symptoms, and tissue inflammation. This work provides a theoretical basis for the rational application of bifenthrin and environmental risk assessment, as well as scientific guidance for the conservation of amphibian populations.

Transient effects of methyltestosterone injection on different reproductive parameters of the hermaphrodite fish Kryptolebias marmoratus.

To elucidate the action mechanism of environmental androgenic chemicals on fish reproductive activity by transient stimulation in heavily polluted areas, individuals of the hermaphrodite fish Kryptolebias marmoratus were injected once with six concentrations of methyltestosterone (MT) (0.1, 1, 5, 10, 50, and 100 µg/g BW) intraperitoneally. The fish were sampled at intervals of 7, 15, and 30 days after a single injection. At 7 days after injection, mature oocytes were not observed in the MT-exposed groups except for the group exposed to 0.1 µg MT, while testicular development was not remarkably different between any of the groups. Also, at 7 days after injection, hepatic estrogen receptor α (ERα) and vitellogenin (VTG) mRNA abundance decreased significantly in the MT-exposed groups despite no significant difference in plasma 17ß-estradiol (E2) levels between any of the groups. This significant difference in VTG mRNA between the control and the MT-exposed groups persisted until 30 days after injection, although ERα mRNA abundance was not statistically different between any groups at 30 days after injection. Our results clearly show that a single injection of MT inhibits ovarian development rather than testicular development in the hermaphroditic gonad of K. marmoratus. Furthermore, our results demonstrate that a single injection of MT interfered with hepatic VTG mRNA synthesis mediated by the suppression of hepatic ERα mRNA transcription.

Chronological Changes in Gonadotropin-Releasing Hormone 1, Gonadotropins, and Sex Steroid Hormones along the Brain-Pituitary-Gonadal Axis during Gonadal Sex Differentiation and Development in the Longtooth Grouper, Epinephelus bruneus.

(1) Fshß and Lhß showed stronger signals and higher transcript levels from 590 to 1050 dph than at earlier stages, implying their active involvement during primary oocyte development. (2) Fshß and Lhß at lower levels were detected during the phases of ovarian differentiation and oogonial proliferation. (3) E2 concentrations increased significantly at 174, 333, and 1435 dph, while T concentrations exhibited significant increases at 174 and 333 dph. These findings suggest potential correlations between serum E2 concentrations and the phases of oogonial proliferation and pre-vitellogenesis.

Male Pheromones Induce Ovulation in Female Honeycomb Groupers (Epinephelus merra): A Comprehensive Study of Spawning Aggregation Behavior and Ovarian Development.

This study characterizes the spawning phenomena of the honeycomb grouper (Epinephelus merra), which is a lunar-synchronized spawner that spawns a few days after full moon. To elucidate the aggregation characteristics of wild honeycomb groupers, the numbers of males and females at the spawning grounds were counted before and after the full moon. Approximately 20 males were consistently observed at the spawning grounds throughout the study period. Females appeared several days after full moon and rapidly increased in number, peaking four days after full moon (41 individuals). The maturation status of the females aggregating at the spawning grounds was investigated. The gonadosomatic index increased rapidly three days after full moon, and ovulation was confirmed. Individuals with ovulatory eggs were present for three days, after which the number of females at the spawning grounds decreased. Additionally, the role of males in final oocyte maturation (FOM) and ovulation in females during the spawning phase was investigated in captivity. FOM was induced in females reared in water with mature males, suggesting that male pheromones in the water induced FOM via activation of the hypothalamic-pituitary-gonadal axis. This suggests that spawning at the natural spawning grounds was the result of male-female interactions via pheromones.

Endocrine Regulation of Maturation and Sex Change in Groupers.

Groupers are widely distributed in tropical and subtropical areas worldwide, are key species to coastal ecosystems, and valuable fishery targets. To facilitate artificial seed production technology for grouper aquaculture, the mechanisms of reproduction and gonad development are being elucidated for these important species. In addition, since groupers are sexually dimorphic fish with female-first maturity (protogynous hermaphrodite fish), research is being conducted to clarify the ecological mechanism of sex change and their reproductive physiology, focusing on the endocrine system. In recent years, research on groupers has also been conducted to understand changes in the coastal environment caused by ocean warming and man-made chemicals. However, due to difficulties associated with conducting research using wild populations for breeding experiments, knowledge of the physiology and ecology of these fish is lacking, especially their reproductive physiology. In this review, we present information on the reproductive physiology and endocrinology of groupers obtained to date, together with the characteristics of their life history.

Marine-derived microbes and molecules for drug discovery.

Increasing attention has been paid to marine-derived biomolecules as sources of therapeutics for autoimmune diseases. Nagasaki Prefecture has many islands and is surrounded by seas, straits, gulfs, bays, and coves, giving it the second longest coastline in Japan after Hokkaido. We have collected more than 20,000 marine microbes and have been preparing an original marine microbial extract library, which contains small and mid-size biomolecules that may penetrate cell membranes and interfere with the intracellular protein-protein interaction involved in the development of autoinflammatory diseases such as familial Mediterranean fever. In addition, we have been developing an indoor shark farming system to prepare shark nanobodies that could be developed as potential therapeutic agents for autoimmune diseases. Sharks produce heavy-chain antibodies, called immunoglobulin new antigen receptors (IgNARs), consisting of one variable domain (VNAR) and five constant domains (CNAR); of these, VNAR can recognize a variety of foreign antigens. A VNAR single domain fragment, called a nanobody, can be expressed in Escherichia coli and has the properties of an ideal therapeutic candidate for autoimmune diseases. Shark nanobodies contain complementarity-determining regions that are formed through the somatic rearrangement of variable, diversity, and joining segments, with the segment end trimming and the N- and P-additions, as found in the variable domains of mammalian antibodies. The affinity and diversity of shark nanobodies are thus expected to be comparable to those of mammalian antibodies. In addition, shark nanobodies are physically robust and can be prepared inexpensively; as such, they may lead to the development of highly specific, stable, effective, and inexpensive biotherapeutics in the future. In this review, we first summarize the history of the development of conventional small molecule drugs and monoclonal antibody therapeutics for autoimmune diseases, and then introduce our drug discovery system at Nagasaki University, including the preparation of an original marine microbial extract library and the development of shark nanobodies.

Dorsal regulates the expression of two phage lysozymes acquired via horizontal gene transfer in triangle sail mussel Hyriopsis cumingii.

Dorsal is a Rel/NF-κB transcription factor, which forms a key part of the Toll pathway. Lysozyme is a ubiquitous enzyme that degrades bacterial cell walls. In this study, a Dorsal homolog was cloned and characterized from triangle sail mussel Hyriopsis cumingii, namely, HcDorsal. Dorsal consisted of 3041 bp, including a 1938 bp open reading frame encoding a 645 amino acid protein. The deduced HcDorsal protein contained a Rel homology domain and an Ig-like, plexin, transcription factor domain. Analysis of expression patterns showed that HcDorsal was highly expressed in the hepatopancreas of H. cumingii. The expression level of HcDorsal continuously increased after Vibrio parahaemolyticus stimulation. When HcDorsal was knocked down by siRNA interference, two phage lysozyme genes (HcLyso1 and HcLyso2) obtained by horizontal gene transfer were significantly downregulated in hemocytes of mussels. Furthermore, knockdown of HcLyso1 and HcLyso2 could weaken V. parahaemolyticus clearance ability. Recombinant HcLyso1 and HcLyso2 proteins accelerated the bacterial clearance in vivo in mussels and evidently inhibited the growth of V. parahaemolyticus. These results suggested that HcDorsal could be activated after V. parahaemolyticus stimulation and then modulate the immune response through the transcriptional regulation of HcLyso1 and HcLyso2, thereby playing a protective role in mussels.