Somatostatin Receptor Gene Functions in Growth Regulation in Bivalve Scallop and Clam.

Bivalves hold an important role in marine aquaculture and the identification of growth-related genes in bivalves could contribute to a better understanding of the mechanism governing their growth, which may benefit high-yielding bivalve breeding. Somatostatin receptor (SSTR) is a conserved negative regulator of growth in vertebrates. Although SSTR genes have been identified in invertebrates, their involvement in growth regulation remains unclear. Here, we identified seven SSTRs (PySSTRs) in the Yesso scallop, Patinopecten yessoensis, which is an economically important bivalve cultured in East Asia. Among the three PySSTRs (PySSTR-1, -2, and -3) expressed in adult tissues, PySSTR-1 showed significantly lower expression in fast-growing scallops than in slow-growing scallops. Then, the function of this gene in growth regulation was evaluated in dwarf surf clams (Mulinia lateralis), a potential model bivalve cultured in the lab, via RNA interference (RNAi) through feeding the clams Escherichia coli containing plasmids expressing double-stranded RNAs (dsRNAs) targeting MlSSTR-1. Suppressing the expression of MlSSTR-1, the homolog of PySSTR-1 in M. lateralis, resulted in a significant increase in shell length, shell width, shell height, soft tissue weight, and muscle weight by 20%, 22%, 20%, 79%, and 92%, respectively. A transcriptome analysis indicated that the up-regulated genes after MlSSTR-1 expression inhibition were significantly enriched in the fat digestion and absorption pathway and the insulin pathway. In summary, we systemically identified the SSTR genes in P. yessoensis and revealed the growth-inhibitory role of SSTR-1 in bivalves. This study indicates the conserved function of somatostatin signaling in growth regulation, and ingesting dsRNA-expressing bacteria is a useful way to verify gene function in bivalves. SSTR-1 is a candidate target for gene editing in bivalves to promote growth and could be used in the breeding of fast-growing bivalves.

Paralytic shellfish toxins producing dinoflagellates cause dysbacteriosis in scallop gut microbial biofilms.

Filter-feeding bivalves could accumulate paralytic shellfish toxins (PSTs) produced by harmful dinoflagellates through diet. Despite that bivalves are resistant to these neurotoxins due to possessing PST-resistant sodium channel, exposure to PSTs-producing dinoflagellates impair bivalve survival. We hypothesized that ingesting PSTs-producing dinoflagellates may influence the gut microbiota, and then the health of bivalves. To test this idea, we compared the gut microbiota of the scallop Patinopecten yessoensis, after feeding with PST-producing or non-toxic dinoflagellates. Exposure to PSTs-producing dinoflagellates resulted in a decline of gut microbial diversity and a disturbance of community structure, accompanied by a significant increase in the abundance and richness of pathogenic bacteria, represented by Vibrio. Moreover, network analysis demonstrated extensive positive correlations between pathogenic bacteria abundances and PSTs concentrations in the digestive glands of the scallops. Furthermore, isolation of a dominant Vibrio strain and its genomic analysis revealed a variety of virulence factors, including the tolC outer membrane exporter, which were expressed in the gut microbiota. Finally, the infection experiment demonstrated scallop mortality caused by the isolated Vibrio strain; further, the pathogenicity of this Vibrio strain was attenuated by a mutation in the tolC gene. Together, these findings demonstrated that the PSTs may affect gut microbiota via direct and taxa-specific interactions with opportunistic pathogens, which proliferate after transition from seawater to the gut environment. The present study has revealed novel mechanisms towards deciphering the puzzles in environmental disturbances-caused death of an important aquaculture species.

Identification, Characterization and Functional Analysis of Fibroblast Growth Factors in Black Rockfish (Sebastes schlegelii).

Fibroblast growth factors (FGFs) are short polypeptides that play essential roles in various cellular biological processes, including cell migration, proliferation, and differentiation, as well as tissue regeneration, immune response, and organogenesis. However, studies focusing on the characterization and function of FGF genes in teleost fishes are still limited. In this study, we identified and characterized expression patterns of 24 FGF genes in various tissues of embryonic and adult specimens of the black rockfish (Sebates schlegelii). Nine FGF genes were found to play essential roles in myoblast differentiation, as well as muscle development and recovery in juvelines of S. schlegelii. Moreover, sex-biased expression pattern of multiple FGF genes was recorded in the species' gonads during its development. Among them, expression of the FGF1 gene was recorded in interstitial and sertoli cells of testes, promoting germ-cell proliferation and differentiation. In sum, the obtained results enabled systematic and functional characterization of FGF genes in S. schlegelii, laying a foundation for further studies on FGF genes in other large teleost fishes.

Establishment of myoblast cell line and identification of key genes regulating myoblast differentiation in a marine teleost, Sebastes schlegelii.

Skeletal myoblasts are activated satellite cells capable of proliferation and differentiation. Studies on mammalian myoblast differentiation and myogenesis could be carried out in vitro thanks to the availability of mouse myoblast cell line C2C12. Lacking of muscle cell line hinders the studies of teleost fish myogenesis. Here, we established a continuous skeletal muscle cell line from juvenile rockfish (Sebastes schlegelii) muscle using explant method and subcultured more than 50 passages for over 150 days. Stable expression of myoblast-specific marker, MyoD (myoblast determination protein) and the potential of differentiation into multi-nucleated skeletal myotubes upon induction suggested the cell line were predominately composed of myoblasts. Transcriptome analysis revealed a total of 4375 genes differentially expressed at four time points after the switch to differentiation medium, which were mainly involved in proliferation and differentiation of myoblasts. KIF22 (kinesin family member 22) and POLA1 (DNA polymerase alpha 1) were identified as the key genes involved in fish myoblast proliferation whereas MYL3 (myosin light chain 3) and TNNT2 (troponin T2) were determined as the crucial genes responsible for differentiation. In all, the continuous myoblasts cultured in this study provided a cell platform for future studies on marine fish myoblast differentiation and myogenesis. The molecular process of myoblast differentiation revealed in this study will open a window into the understanding of indeterminate muscle growth of large teleost.

Genome-Wide Identification, Characterization and Expression Profiling of myosin Family Genes in Sebastes schlegelii.

Myosins are important eukaryotic motor proteins that bind actin and utilize the energy of ATP hydrolysis to perform a broad range of functions such as muscle contraction, cell migration, cytokinesis, and intracellular trafficking. However, the characterization and function of myosin is poorly studied in teleost fish. In this study, we identified 60 myosin family genes in a marine teleost, black rockfish (Sebastes schlegelii), and further characterized their expression patterns. myosin showed divergent expression patterns in adult tissues, indicating they are involved in different types and compositions of muscle fibers. Among 12 subfamilies, S. schlegelii myo2 subfamily was significantly expanded, which was driven by tandem duplication events. The up-regulation of five representative genes of myo2 in the skeletal muscle during fast-growth stages of juvenile and adult S. schlegelii revealed their active role in skeletal muscle fiber synthesis. Moreover, the expression regulation of myosin during the process of myoblast differentiation in vitro suggested that they contribute to skeletal muscle growth by involvement of both myoblast proliferation and differentiation. Taken together, our work characterized myosin genes systemically and demonstrated their diverse functions in a marine teleost species. This lays foundation for the further studies of muscle growth regulation and molecular mechanisms of indeterminate skeletal muscle growth of large teleost fishes.

A chromosome-level genome of black rockfish, Sebastes schlegelii, provides insights into the evolution of live birth.

The black rockfish (Sebastes schlegelii) is a teleost in which eggs are fertilized internally and retained in the maternal reproductive system, where they undergo development until live birth (viviparity). In the present study, we report a chromosome-level black rockfish genome assembly. High-throughput transcriptome analysis (RNA-seq and ATAC-seq) coupled with in situ hybridization (ISH) and immunofluorescence reveal several candidate genes for maternal preparation, sperm storage and release, and hatching. We propose that zona pellucida (ZP) proteins retain sperm at the oocyte envelope, while genes in two distinct astacin metalloproteinase subfamilies serve to release sperm from the ZP and free the embryo from chorion at prehatching stage. We present a model of black rockfish reproduction, and propose that the rockfish ovarian wall has a similar function to the uterus of mammals. Together, these genomic data reveal unprecedented insights into the evolution of an unusual teleost life history strategy, and provide a sound foundation for studying viviparity in nonmammalian vertebrates and an invaluable resource for rockfish ecological and evolutionary research.

Mangiferin Inhibits Renal Urate Reabsorption by Modulating Urate Transporters in Experimental Hyperuricemia.

Mangiferin, a natural glucosyl xanthone from the leaves of Mangifera indica L., was previously shown to exert potent hypouricemic effects associated with inhibition of the activity of xanthine dehydrogenase/oxidase. The present study aimed to evaluate its uricosuric effect and possible molecular mechanisms underlying the renal urate transporters responsible for urate reabsorption in vivo. Mangiferin (1.5-24.0 mg/kg) was administered intragastrically to hyperuricemic mice and rats induced by the intraperitoneal injection of uric acid and potassium oxonate, respectively. The uricosuric effect was evaluated by determining the serum and urinary urate levels as well as fractional excretion of uric acid (FEUA). The mRNA and protein levels of renal urate-anion transporter 1 (URAT1), organic anion transporter 10 (OAT10), glucose transporter 9 (GLUT9), and PDZ domain-containing protein (PDZK1) were analyzed. The administration of mangiferin significantly decreased the serum urate levels in hyperuricemic mice in a dose- and time-dependent manner. In hyperuricemic rats, mangiferin also reduced the serum urate levels and increased the urinary urate levels and FEUA. These results indicate that mangiferin has uricosuric effects. Further examination showed that mangiferin markedly inhibited the mRNA and protein expression of renal URAT1, OAT10, and GLUT9 in hyperuricemic rats, but did not interfere with PDZK1 expression. Taken together, these findings suggest that mangiferin promotes urate excretion by the kidney, which may be related to the inhibition of urate reabsorption via downregulation of renal urate transporters.