RESUMO
In order to comprehend the molecular basis of growth, nutrient composition, and color pigmentation in oysters, comparative proteome and metabolome analyses of two selectively bred oyster strains with contrasting growth rate and shell color were used in this study. A total of 289 proteins and 224 metabolites were identified differentially expressed between the two strains. We identified a series of specifically enriched functional clusters implicated in protein biosynthesis (RPL4, MRPS7, and CARS), fatty acid metabolism (ACSL5, PEX3, ACOXI, CPTIA, FABP6, and HSD17B12), energy metabolism (FH, PPP1R7, CLAM2, and RGN), cell proliferation (MYB, NFYC, DOHH, TOP2a, SMARCA5, and SMARCC2), material transport (ABCB1, ABCB8, VPS16, and VPS33a), and pigmentation (RDH7, RDH13, Retsat, COX15, and Cyp3a9). Integrated proteome and metabolome analyses indicate that fast-growing strain utilize energy-efficient mechanisms of ATP generation while promoting protein and polyunsaturated fatty acid synthesis, activating the cell cycle to increase cell proliferation and thus promoting their biomass increase. These results uncovered molecular mechanisms underlying growth regulation, nutrition quality, and pigmentation and provided candidate biomarkers for molecular breeding in oysters. SIGNIFICANCE: Rapid growth has always been the primary breeding objective to increase the production profits of Pacific oyster (Crassostrea gigas), while favorable nutritional quality and beautiful color add commercial value. In recent years, proteomic and metabolomic techniques have been widely used in marine organisms, although these techniques are seldom utilized to study oyster growth and development. In this study, two C. gigas strains with contrasted phenotypes in growth and shell color provided an ideal model for unraveling the molecular basis of growth and nutrient composition through a comparison of the proteome and metabolome. Since proteins and metabolites are the critical undertakers and the end products of cellular regulatory processes, identifying the differentially expressed proteins and metabolites would allow for discovering biomarkers and pathways that were implicated in cell growth, proliferation, and other critical functions. This work provides valuable resources in assistance with molecular breeding of oyster strains with superior production traits of fast-growth and high-quality nutrient value.
Assuntos
Crassostrea , Animais , Crassostrea/metabolismo , Proteoma/metabolismo , Proteômica/métodos , Metaboloma , Nutrientes , Biomarcadores/metabolismoRESUMO
Parallel diversification provides a proper framework for studying the role of natural selection in evolution. Yet, empirical studies from ecological 'non-model' species of invertebrates are limited at the whole genome level. Here, we presented a chromosome-scale genome assembly for Crassostrea angulata and investigated the parallel genomic evolution in oysters. Specifically, we used population genomics approaches to compare two southern-northern oyster species pairs (C. angulata-C. gigas and southern-northern C. ariakensis) along the coast of China. The estimated divergence time of C. angulata and C. gigas is earlier than that of southern and northern C. ariakensis, which aligns with the overall elevated genome-wide divergence. However, the southern-northern C. ariakensis FST profile represented more extremely divergent "islands". Combined with recent reciprocal hybridization studies, we proposed that they are currently at an early stage of speciation. These two southern-northern oyster species pairs exhibited significant repeatability in patterns of genome-wide differentiation, especially in genomic regions with extremely high and low divergence. This suggested that divergent and purifying selection has contributed to the genomic parallelism between southern and northern latitudes. Top differentiated genomic regions shared in these two oyster species pairs contained candidate genes enriched for functions in energy metabolism, especially adipogenesis, which are closely related to reproductive behaviours. These genes might be good candidates for further investigation in vivo. In conclusion, our results suggest that similar divergent selection and shared genomic features could predictably transform standing genetic variation within one species pair into differences in another.
Assuntos
Crassostrea , Animais , Crassostrea/genética , Adipogenia/genética , Genoma/genética , Cromossomos , ChinaRESUMO
The Pacific oyster (Crassostrea gigas) is a widely cultivated shellfish in the world, while its transcriptome diversity remains less unexplored due to the limitation of short reads. In this study, we used Oxford Nanopore sequencing to develop the full-length transcriptome database of C. gigas. We identified 77,920 full-length transcripts from 21,523 genes, and uncovered 9668 alternative splicing events and 87,468 alternative polyadenylation sites. Notably, a total of 16,721 novel transcripts were annotated in this work. Furthermore, integrative analysis of 25 publicly available RNA-seq datasets revealed the transcriptome diversity involved in post-transcriptional regulation in C. gigas. We further developed a Drupal based webserver, Cgtdb, which can be used for transcriptome visualization, sequence alignment, and functional genome annotation analyses. This work provides valuable resources and a useful tool for integrative analysis of various transcriptome datasets in C. gigas, which will serve as an essential reference for functional annotation of the oyster genome.
RESUMO
Ocean temperature rising drastically threatens the adaptation and survival of marine organisms, causing serious ecological impacts and economic losses. It is crucial to understand the adaptive mechanisms of marine organisms in response to high temperature. In this study, a novel regulatory mechanism that is mediated by hypoxia-inducible factor-1α (HIF-1α) was revealed in Pacific oyster (Crassostrea gigas) in response to heat stress. We identified a total of six HIF-1α genes in the C. gigas genome, of which HIF-1α and HIF-1α-like5 were highly induced under heat stress. We found that the HIF-1α and HIF-1α-like5 genes played critical roles in the heat shock response (HSR) through upregulating the expression of heat shock protein (HSP). Knocking down of HIF-1α via RNA interference (RNAi) inhibited the expression of heat shock factor 1 (HSF1) and HSP70 genes in C. gigas under heat stress. Both HIF-1α and HIF-1α-like5 promoted the transcriptional activity of HSF1 by binding to hypoxia response elements (HREs) within the promoter region. Furthermore, the survival of C. gigas under heat stress was significantly decreased after knocking down of HIF-1α. This work for the first time revealed the involvement of HIF-1α/HSF1/HSP70 pathway in response to heat stress in the oyster and provided an insight into adaptive mechanism of bivalves in the face of ocean warming.
Assuntos
Crassostrea , Animais , Crassostrea/metabolismo , Regulação da Expressão Gênica , Resposta ao Choque Térmico/genética , Proteínas de Choque Térmico HSP70/genética , Proteínas de Choque Térmico HSP70/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Transdução de SinaisRESUMO
Marine ecosystems with ocean warming and industry pollution threaten the survival and adaptation of organisms. G protein-coupled receptors (GPCRs) play critical roles in various physiological and toxicological processes in vertebrates and invertebrates. The Pacific oyster (Crassostrea gigas) was widely used to study the adaptation of marine molluscs to coastal environments. In this work, we identified a total of 586 GPCRs in C. gigas genome. The C. gigas GPCRs were divided into five classes (including class A, B, C, E and F) with different degrees of expansion. Meta-analysis of multiple RNA-seq datasets revealed that transcriptional expression patterns of GPCRs in C. gigas were distinct in response to high temperature, salinity, air exposure, heavy metal, ostreid herpes virus 1 (OsHV-1) and Vibrio challenge. This work for the first time characterized the GPCR gene family and provided insights into the potential roles of GPCRs in adaptation of marine molluscs to stressful coastal environment.
Assuntos
Crassostrea , Animais , Crassostrea/metabolismo , Duplicação Gênica , Ecossistema , Salinidade , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismoRESUMO
Fast growth is one of the most important breeding goals for all economic species such as the Pacific oyster (Crassostrea gigas), an aquaculture mollusk with top global production. Although the genetic basis and molecular mechanisms of growth-related traits have been widely investigated in the oyster, the role of DNA methylation involved in growth regulation remains largely unexplored. In this study, we performed a comparative DNA methylome analysis of two selectively bred C. gigas strains with contrasted phenotypes in growth and shell color based on whole-genome bisulfite sequencing (WGBS). Genome-wide profiling of DNA methylation at the single-base resolution revealed that DNA methylations were widely spread across the genome with obvious hotspots, coinciding with the distribution of genes and repetitive elements. Higher methylation levels were observed within genic regions compared with intergenic and promoter regions. Comparative analysis of DNA methylation allowed the identification of 339,604 differentially methylated CpG sites (DMCs) clustering in 27,600 differentially methylated regions (DMRs). Functional annotation analysis identified 11,033 genes from DMRs which were enriched in biological processes including cytoskeleton system, cell cycle, signal transduction, and protein biosynthesis. Integrative analysis of methylome and transcriptome profiles revealed a positive correlation between gene expression and DNA methylation within gene-body regions. Protein-protein interaction (PPI) analysis of differentially expressed and methylated genes allowed for the identification of integrin beta-6 (homolog of human ITGB3) as a hub modulator of the PI3K/Akt signaling pathway that was involved in various growth-related processes. This work provided insights into epigenetic regulation of growth in oysters and will be valuable resources for studying DNA methylation in invertebrates.
Assuntos
Fenômenos Biológicos , Crassostrea , Animais , Metilação de DNA , Epigênese Genética , Epigenoma , Humanos , Integrinas/genética , Integrinas/metabolismo , Fosfatidilinositol 3-Quinases/genética , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-akt/metabolismoRESUMO
Neuroendocrine hormones such as dopamine and insulin/insulin-like peptides play indispensable roles in growth regulation of animals, while the interplay between dopamine and insulin signaling pathways remains largely unknown in invertebrates. In the present study, we showed that tyrosine hydroxylase (TH), the rate-limiting enzyme of dopamine synthesis, was highly expressed in all tissues of the fast-growing oysters, and gradually increased with the development, which indicated the potential role of dopamine in growth regulation. Incubated with dopamine hydrochloride and insulin-like peptide recombinant proteins in vitro induced the expression of TH, suggesting a mutual regulatory relationship between insulin and dopamine signaling. Fasting and re-feeding experiments confirmed the role of TH in food intake regulation, also provide a clue about the potential regulatory relationship between the FoxO and TH. Further luciferase assay experiment confirmed that FoxO was involved in transcriptional regulation of TH gene through binding to its specific promoter region. This work provided insights into the crosstalk between dopamine and insulin signaling in growth control of mollusks.
Assuntos
Dopamina , Ostreidae , Animais , Dopamina/metabolismo , Insulina/metabolismo , Ostreidae/metabolismo , Transdução de Sinais , Tirosina 3-Mono-Oxigenase/metabolismoRESUMO
Cadmium (Cd) is one of the most harmful heavy metals due to its persistence and bioaccumulation through the food chains, posing health risks to human. Oysters can bioaccumulate and tolerate high concentrations of Cd, providing a great model for studying molecular mechanism of Cd detoxification. In a previous study, we identified two CYP genes, CYP17A1-like and CYP2C50, that were potentially involved in Cd detoxification in the Pacific oyster, Crassostrea gigas. In this work, we performed further investigations on their physiological roles in Cd detoxification through RNA interference (RNAi). After injection of double-stranded RNA (dsRNA) into the adductor muscle of oysters followed by Cd exposure for 7 days, we observed that the expressions of CYP17A1-like and CYP2C50 in interference group were significantly suppressed on day 3 compared with control group injected with PBS. Moreover, the mortality rate and Cd content in the CYP17A1-like dsRNA interference group (dsCYP17A1-like) was significantly higher than those of the control on day 3. Furthermore, the activities of antioxidant enzymes, including SOD, CAT, GST, were significantly increased in dsCYP17A1-like group, while were not changed in dsCYP2C50 group. More significant tissue damage was observed in gill and digestive gland of oysters in RNAi group than control group, demonstrating the critical role of CYP17A1-like in Cd detoxification. Dual luciferase reporter assay revealed three core regulatory elements of MTF-1 within promoter region of CYP17A1-like, suggesting the potential transcriptional regulation of CYP17A1-like by MTF-1 in oysters. This work demonstrated a critical role of CYP17A1-like in Cd detoxification in C. gigas and provided a new perspective toward unravelling detoxification mechanisms of bivalves under heavy metal stress.
Assuntos
Crassostrea , Poluentes Químicos da Água , Animais , Cádmio/metabolismo , Cádmio/toxicidade , Crassostrea/genética , Crassostrea/metabolismo , Regulação da Expressão Gênica , Brânquias/metabolismo , Poluentes Químicos da Água/toxicidadeRESUMO
The involvement of insulin/insulin-like growth factor signaling (IIS) pathways in the growth regulation of marine invertebrates remains largely unexplored. In this study, we used a fast-growing Pacific oyster (Crassostrea gigas) variety "Haida No.1" as the material with which to unravel the role of IIS systems in growth regulation in oysters. Systematic bioinformatics analyses allowed us to identify major components of the IIS signaling pathway and insulin-like peptide receptor (ILPR)-mediated signaling pathways, including PI3K-AKT, RAS-MAPK, and TOR, in C. gigas. The expression levels of the major genes in IIS and its downstream signaling pathways were significantly higher in "Haida No.1" than in wild oysters, suggesting their involvement in the growth regulation of C. gigas. The expression profiles of IIS and its downstream signaling pathway genes were significantly altered by nutrient abundance and culture temperature. These results suggest that the IIS signaling pathway coupled with the ILPR-mediated signaling pathways orchestrate the regulation of energy metabolism to control growth in Pacific oysters.
Assuntos
Crassostrea/genética , Receptores de Peptídeos/metabolismo , Somatomedinas/metabolismo , Animais , Biologia Computacional/métodos , Expressão Gênica/genética , Perfilação da Expressão Gênica/métodos , Insulina/metabolismo , Sistema de Sinalização das MAP Quinases/fisiologia , Peptídeos/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptores Proteína Tirosina Quinases/metabolismo , Transdução de Sinais/genética , Somatomedinas/fisiologia , Transcriptoma/genéticaRESUMO
Transmembrane proteins are involved in an array of stress responses, particularly in thermo-sensation and thermo-regulation. In this study, we performed a genome-wide identification and characterization of the Transient Receptor Potential (TRP) genes in the Pacific oyster (Crassostrea gigas) and investigated their expression profiles after heat stress to identify critical TRPs potentially associated with thermal regulation. A total of 66 TRP genes were identified in the C. gigas, which showed significant gene expansion and tandem duplication. Meta-analysis of the available RNA-Seq data generated from samples after acute heat stress revealed a set of heat-inducible TRPs. Further examination of their expression profiles under chronic heat stress, and comparison between C. gigas and C. angulata, two oyster species with different tolerance levels to heat stress, led to the identification of TRPC3.6, TRPC3.7, and TRPV4.7 as important TRPs involved in thermal regulation in oysters. This work provided valuable information for future studies on the molecular mechanism of TRP mediated thermal tolerance, and identification of diagnostic biomarker for thermal stress in the oysters.
Assuntos
Crassostrea/fisiologia , Resposta ao Choque Térmico/genética , Transcriptoma , Canais de Potencial de Receptor Transitório/genética , Animais , Biologia Computacional/métodos , Crassostrea/classificação , Dosagem de Genes , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Estudo de Associação Genômica Ampla , Invertebrados , Fenótipo , Filogenia , Estresse Fisiológico/genética , VertebradosRESUMO
The insulin/insulin-like growth factor signaling (IIS) pathway is well-known in regulation of cell growth and proliferation in vertebrates, while its role in invertebrates such as mollusks remains largely unknown. In this study, we performed an extensive multi-omics data mining and identified four insulin-like peptide genes, including ILP, MIRP3, MIRP3-like and ILP7, in the Pacific oyster, Crassostrea gigas. Their potential roles in growth regulation were further investigated using the selectively bred fast-growing C. gigas variety "Haida No.1". Expression profiling and in situ hybridization of these insulin-like peptides suggested their distinct tissue-specific expression pattern, with dominant expression in the neural enrichment tissues such as labial palp, visceral ganglia, adductor muscle, and digestive gland. The expressions of insulin-like peptides were significantly altered by food abundance in a gene-specific fashion. The expression of ILP was reduced during fasting and increased after re-feeding, the expressions of MIRP3 and ILP7 were generally induced during fasting and down-regulated after re-feeding, while the expression of MIRP3-like was firstly up-regulated and then down-regulated during the fasting and re-feeding process. Furthermore, the expressions of all four insulin-like peptide genes were significantly suppressed at low temperature, in accordance with the growth inhibition. These results indicated that all four insulin-like peptides would play critical but different roles in regulation of growth in the oysters. This work provides valuable information for further investigation on growth regulation mechanism in mollusks and molecular assisted breeding of growth with other production traits in the Pacific oyster.
Assuntos
Crassostrea/crescimento & desenvolvimento , Crassostrea/metabolismo , Perfilação da Expressão Gênica , Insulina/metabolismo , Peptídeos/metabolismo , Sequência de Aminoácidos , Animais , Insulina/genética , Peptídeos/genética , FilogeniaRESUMO
Fast growth is one of the most desired traits for all food animals, which affects the profitability of animal production. The Pacific oyster, Crassostrea gigas, is an important aquaculture shellfish around the world with the largest annual production. Growth of the Pacific oyster has been greatly improved by artificial selection breeding, but molecular mechanisms underlying growth remains poorly understood, which limited the molecular integrative breeding of fast growth with other superior traits. In this study, comparative transcriptome analyses between the fast-growing selectively bred Pacific oyster and unselected wild Pacific oysters were conducted by RNA-Seq. A total of 1,303 protein-coding genes differentially expressed between fast-growing oysters and wild controls were identified, of which 888 genes were expressed at higher levels in the fast-growing oysters. Functional analysis of the differentially expressed genes (DEGs) indicated that genes involved in microtubule motor activity and biosynthesis of nucleotides and proteins are potentially important for growth in the oyster. Positive selection analysis of genes at the transcriptome level showed that a significant number of ribosomal protein genes had undergone positive selection during the artificial selection breeding process. These results also indicated the importance of protein biosynthesis and metabolism for the growth of oysters. The alternative splicing (AS) of genes was also compared between the two groups of oysters. A total of 3,230 differential alternative splicing events (DAS) were identified, involved in 1,818 genes. These DAS genes were associated with specific functional pathways related to growth, such as "long-term potentiation," "salivary secretion," and "phosphatidylinositol signaling system." The findings of this study will be valuable resources for future investigation to unravel molecular mechanisms underlying growth regulation in the oyster and other marine invertebrates and to provide solid support for breeding application to integrate fast growth with other superior traits in the Pacific oyster.