Characterization of a pseudohemocyanin gene (PtPhc1) and its immunity function in response to Vibrio parahaemolyticus infection in the swimming crab Portunus trituberculatus.

Pseudohemocyanin is a member of the hemocyanin superfamily, but little research is available on its function in immunology. In this study, a Portunus trituberculatus pseudohemocyanin gene, named PtPhc1, was obtained by gene cloning. The PtPhc1 cDNA was 2312 bp in length, encoding 684 amino acids while exhibiting a characteristic hemocyanin structural domain. Tissue expression analysis revealed ubiquitous expression of PtPhc1 across all tissues, with the highest level of expression observed in the hepatopancreas. The expression pattern of PtPhc1 in response to Vibrio parahaemolyticus infection was clarified using RT-qPCR in swimming crabs. Notably, the expression peaked at 24 h, and increased 1435-fold compared to the control group in the hepatopancreas. While the expression level reached the maximum value at 72 h, which was 3.24 times higher than that of the control group in hemocytes. Remarkably, the reduction in PtPhc1 expression led to a noteworthy 30% increase in the mortality rate of P. trituberculatus when exposed to V. parahaemolyticus. In addition, in vitro bacterial inhibition assays exhibited a dose-dependent suppression of bacterial proliferation by recombinant PtPhc1 protein, with a notable inhibition rate of 48.33% against V. parahaemolyticus at a concentration of 0.03 mg/mL. To the best of our knowledge, the results establish the function of pseudohaemocyanin in immunity for the first time, contributing to a deeper comprehension of innate immune regulatory mechanisms in aquatic organisms and advancing strategies for disease-resistant breeding.

A chromosome-level genome of the kuruma shrimp (Marsupenaeus japonicus) provides insights into its evolution and cold-resistance mechanism.

Marsupenaeus japonicus is an important marine crustacean species. However, a lack of genomic resources hinders the use of whole genome sequencing to explore their genetic basis and molecular mechanisms for genome-assisted breeding. Consequently, we determined the chromosome-level genome of M. japonicus. Here we determine the chromosome-level genome assembly for M. japonicus with a total of 665.19 Gb genomic sequencing data, yielding an approximately1.54 Gb assembly with a contig N50 size of 229.97 kb and a scaffold N50 size of 38.27 Mb. With the high-throughput chromosome conformation capture (Hi-C) technology, we anchored 18,019 contigs onto 42 pseudo-chromosomes, accounting for 99.40% of the total genome assembly. Analysis of the present M. japonicus genome revealed 24,317 protein-coding genes and a high proportion of repetitive sequences (61.56%). The high-quality genome assembly enabled the identification of genes associated with cold-stress and cold tolerance in kuruma shrimp through the comparison of eyestalk transcriptomes between the low temperature-stressed shrimp (10 °C) and normal temperature shrimp (28 °C). The genome assembly presented here could be useful in future studies to reveal the molecular mechanisms of M. japonicus in response to low temperature stress and the molecular assisted breeding of M. japonicus in low temperature.

QTL mapping and marker identification for sex determination in the ridgetail white prawn, Exopalaemon carinicauda.

Sex determination is an important and intriguing research topic in the field of evolutionary and developmental biology. Quantitative trait locus (QTL) mapping for sex is helpful in clarifying the sex determination system of species. In this study, a second high-resolution genetic linkage map was constructed for the ridgetail white prawn, Exopalaemon carinicauda, which included 9280 markers, covering 99.98% of the complete genome. Based on the linkage map, a highly significant sex-related QTL was first mapped to a single linkage group (LG3, LOD > 55.6). Fifty-two markers in the QTL region were significantly associated with sex (p ≤ 10-40), of which heterogametic genotypes in females supported the ZW sex determination mechanism. Six markers were verified to be significantly associated with sex in the wild population. Some sex-related genes were identified, including phospholipase D, protein kinase shaggy, and longitudinals lacking protein. These results inform our understanding of the mechanisms of sex determination in E. carinicauda.

Genome survey and high-resolution backcross genetic linkage map construction of the ridgetail white prawn Exopalaemon carinicauda applications to QTL mapping of growth traits.

BACKGROUND: High-resolution genetic linkage map is critical for QTL mapping, genome sequence assembly and marker-assisted selection in aquaculture species. The ridgetail white prawn Exopalaemon carinicauda is one of the most economic shrimp species naturally distributed in the coasts of eastern China and western Korea. However, quite limited genomics and genetics information have been exploited for genetic improvement of economic traits in this species. RESULTS: In the present study, we conducted genome survey and constructed high-resolution genetic linkage maps of the ridgetail white prawn with reciprocal-cross mapping family genotyped using next-generation sequencing approaches. The estimated genome size was 9.33 Gb with a heterozygosity of 0.26% and a repeat sequence ratio of 76.62%. 65,772 protein-coding genes were identified by genome annotation. A total of 10,384 SNPs were used to high-throughput genotyping and assigned to 45 linkage groups (LGs) from reciprocal backcross families of E. carinicauda, and the average marker distances were 0.73 cM and 0.55 cM, respectively. Based on the high-resolution linkage map, twenty-three QTLs related to five growth traits were detected. All QTLs could explain 8.8-15.7% of the total growth-traits variation. CONCLUSIONS: The genome size of E. carinicauda was estimated more accurately by genome survey analysis, which revealed basic genomic architecture. The first high-resolution backcross genetic linkage map and QTLs related to growth traits will provide important information for QTL fine mapping, genome assembly and genetic improvement of E. carinicauda and other palaemon shrimps.

Immune response and antioxidant status of Portunus trituberculatus inoculated with pathogens.

The white spot syndrome virus (WSSV), Vibrio parahaemolyticus and V. alginolyticus are serious epidemic pathogen affecting the cultured Portunus trituberculatus and resulted in severe economic losses to local farmers. The immune and antioxidant systems are believed to be closely involved in host responses to pathogens in aerobic animals, including crustaceans. In order to explore such host-pathogen interactions in the early stage of infection in P. trituberculatus, the mRNA transcript levels of six key immune-related genes (proPO, α2M, crustin, lysozyme, NOS, and NOX) and three key antioxidant-related genes (CuZnSOD, CAT, and GPx) and their corresponding enzymatic activity were investigated in response to challenge with the three pathogens. A decrease in the expression of the proPO, crustin, and lysozyme was observed, which may reflect the immunosuppressive mechanism of the pathogens against the host. Moreover, an increase was observed in the α2M expression with time, which indicated that the pathogens could affect proteinase cascade reactions associated with the proPO system by disturbing the balance between serine proteinases and their inhibitors. Moreover, WSSV, V. parahaemolyticus and V. alginolyticus induced to increase the transcription and enzyme activities of NOS and NOX. Additionally, significant variations in the expression of the anitioxidant-related genes CuZnSOD, CAT, and GPx and their enzyme activities implied that these enzymes played a critical role in the immune response against the pathogens. The present findings indicate that the immune parameters analyzed here may be markers of the physiological status of this species after bacterial or viral infections.

Chromosome-level genome assembly of ridgetail white shrimp Exopalaemon carinicauda.

Exopalaemon carinicauda, a eurythermal and euryhaline shrimp, contributes one third of the total biomass production of polyculture ponds in eastern China and is considered as a potential ideal experimental animal for research on crustaceans. We conducted a high-quality chromosome-level genome assembly of E. carinicauda combining PacBio HiFi and Hi-C sequencing data. The total assembly size was 5.86 Gb, with a contig N50 of 235.52 kb and a scaffold N50 of 138.24 Mb. Approximately 95.29% of the assembled sequences were anchored onto 45 pseudochromosomes. BUSCO analysis revealed that 92.89% of 1,013 single-copy genes were highly conserved orthologs. A total of 44, 288 protein-coding genes were predicted, of which 70.53% were functionally annotated. Given its high heterozygosity (2.62%) and large proportion of repeat sequences (71.49%), it is one of the most complex genome assemblies. This chromosome-scale genome will be a valuable resource for future molecular breeding and functional genomics research on E. carinicauda.

Chromosome-level genome of the long-tailed marine-living ornate spiny lobster, Panulirus ornatus.

Recent conservation efforts to protect rare and endangered aquatic species have intensified. Nevertheless, the ornate spiny lobster (Panulirus ornatus), which is prevalent in the Indo-Pacific waters, has been largely ignored. In the absence of a detailed genomic reference, the conservation and population genetics of this crustacean are poorly understood. Here, We assembled a comprehensive chromosome-level genome for P. ornatus. This genome-among the most detailed for lobsters-spans 2.65 Gb with a contig N50 of 51.05 Mb, and 99.11% of the sequences with incorporated to 73 chromosomes. The ornate spiny lobster genome comprises 65.67% repeat sequences and 22,752 protein-coding genes with 99.20% of the genes functionally annotated. The assembly of the P. ornatus genome provides valuable insights into comparative crustacean genomics and endangered species conservation, and lays the groundwork for future research on the speciation, ecology, and evolution of the ornate spiny lobster.

Comparative Transcriptome Analysis of the Response to Vibrio parahaemolyticus and Low-Salinity Stress in the Swimming Crab Portunus trituberculatus.

Vibrio parahaemolyticus is one of the main pathogenic bacteria of Portunus trituberculatus and causes mass mortality of P. trituberculatus in aquaculture. In addition, low-salinity stimulation makes P. trituberculatus more susceptible to V. parahaemolyticus infections. In order to elucidate the molecular mechanism of resistance to V. parahaemolyticus in P. trituberculatus, comparative transcriptomic analysis of blood cells stimulated by low salinity and V. parahaemolyticus was carried out in this study. Transcriptome sequencing of low-salinity stress and pathogen infection at different time points was completed using Illumina sequencing technology. A total of 5827, 6432, 5362 and 1784 differentially expressed genes (DEGs) involved in pathways related to ion transport and immunoregulation were found under low-salinity stress at 12, 24, 48 and 72 h compared with the control at 0 h. In contrast, 4854, 4814, 5535 and 6051 DEGs, which were significantly enriched in Toll and IMD signaling pathways, were found at 12, 24, 48 and 72 h compared with the control at 0 h under V. parahaemolyticus infection. Among them, 952 DEGs were shared in the two treatment groups, which were mainly involved in apoptosis and Hippo signaling pathway. Cluster analysis screened 103 genes that were differentially expressed in two factors that were negatively correlated, including immunoglobulin, leukocyte receptor cluster family, scavenger receptor, macroglobulin and other innate-immune-related genes. These results provide data support for the analysis of the mechanisms of immunity to V. parahaemolyticus under low-salinity stress in P. trituberculatus and help to elucidate the molecular mechanisms by which environmental factors affect immunity.

Molecular Characterization Related to Ovary Early Development Mechanisms after Eyestalk Ablation in Exopalaemon carinicauda.

Eyestalk ablation is an effective method to promote ovarian development in crustaceans. Herein, we performed transcriptome sequencing of ovary and hepatopancreas tissues after eyestalk ablation in Exopalaemon carinicauda to identify genes related to ovarian development. Our analyses led to the identification of 97,383 unigenes and 190,757 transcripts, with an average N50 length of 1757 bp. In the ovary, four pathways related to oogenesis and three related to oocyte rapid growth were enriched. In the hepatopancreas, two vitellogenesis-associated transcripts were identified. Furthermore, short time-series expression miner (STEM) and gene ontology (GO) enrichment analyses revealed five terms related to gamete generation. In addition, two-color fluorescent in situ hybridization results suggested that dmrt1 might play a vital role in oogenesis during the early stage of ovarian development. Overall, our insights should support future studies focusing on investigating oogenesis and ovarian development in E. carinicauda.

Ammonia Stress Disturbs Moult Signaling in Juvenile Swimming Crab Portunus trituberculatus.

Ammonia is a significant concern during hatchery culture in brachyuran species, and its accumulation may lead to abortive moulting and large-scale deaths of the early juveniles. To date, the underlying mechanism for ammonia-induced alteration of the moulting process is still unknown. In this study, we aimed to investigate the effects of ammonia on the moulting as well as the potential mechanisms in early juveniles of the swimming crab Portunus trituberculatus, an important aquaculture species in China. We evaluated the survival rate and moulting rate of the juvenile crabs (C2) and analyzed the expression pattern of the genes in key components of molt signaling during a complete moulting cycle under different concentrations of ammonia nitrogen (the control group: <0.1 mg/L; the LA group: 5 mg/L; and the HA group: 20 mg/L). The results showed that: (1) the survival rate in the LA and HA groups was lower than that in the control group at the end of the experiment, and moulting death syndrome (MDS) was only observed in the HA group; (2) the moulting rate was higher in the LA group and lower in the HA group compared to the control group; (3) consistent with the results of the moulting experiment, MIH showed decreased expression, and genes related to ecdysteroid synthesis, ecdysteroid receptors, and responsive effectors exhibited increased expression in the LA group compared to the control group; and (4) although MIH expression was upregulated, increased expression of the genes associated with ecdysteroid synthesis, ecdysteroid receptors and downstream effectors still observed in the HA group. Our results indicated that low levels of ammonia can promote moulting in juvenile swimming crabs by inhibiting the expression of MIH and activating moult signaling, whereas high levels of ammonia inhibit moulting and lead to MDS through impairing moult signaling.

Identification, characterization and the interaction of Tollip and IRAK-1 in grass carp (Ctenopharyngodon idellus).

Tollip and IRAK-1 are key components of the TLR/IL-1R signaling pathway in mammals, which play crucial roles as mediators of the TLR/IL-1R signal transduction pathways. Although several TLRs have been found in fish, molecular associations, protein-protein interactions or the role of the TLR signaling pathway in infection-induced immunity in fish has received little attention. In this study, Tollip and IRAK-1 sequences of grass carp were isolated from a head kidney cDNA library. Full length transcripts and sequences of promoter regions were obtained by 3' and 5' RACE and genome walking, respectively. Reporter gene-promoter constructs and real-time RT-PCR analysis was used to determine grass carp Tollip and IRAK-1 transcription pattern in tissues. Recombinant proteins were used for antibodies production. Phylogenetically, the grass carp loci clustered with previously reported Tollip and IRAK-1genes, respectively, and their sequences shared the highest identity with the genes of zebrafish (Danio rerio). The promoter region of grass carp Tollip and IRAK-1 proved to be active. After viral infection transcript levels of both loci were upregulated in most immune-related tissues in a time-dependent manner. Using antibodies produced in this study, immunofluorescence analysis indicated that Tollip and IRAK-1 were uniformly distributed and co-localized in the cytoplasm of CIK cells. After viral infection, however, Tollip and IRAK-1 both trended toward the cell membrane. Our results demonstrate the existence of Tollip and IRAK-1 proteins in teleost species, and suggest that Tollip-IRAK-1 complexes are being recruited to receptor complexes after stimulation with virus. These results provide novel insights into the role of the TLR signaling pathway in teleosts, especially the action of teleost Tollip and IRAK-1 and the interaction of these molecules as part of this pathway.

Cloning and characterization of the grass carp (Ctenopharyngodon idella) Toll-like receptor 22 gene, a fish-specific gene.

Toll-like receptor 22 (TLR22) is a fish-specific TLR which recognizes double-strand (ds) RNA and participates in the innate immune response through the Toll-IL-1R homology domain-containing adaptor protein 1 (TICAM-1). To further investigate how the innate immune system of teleosts responds to viral infections, we cloned the full-length cDNA sequence of grass carp (Ctenopharyngodon idella) TLR22 (CiTLR22). The complete cDNA sequence of CiTLR22 was 3647 bp and encodes a polypeptide of 954 amino acids. Analysis of the deduced amino acid sequence indicated that CiTLR22 has typical structural features of proteins belonging to the TLR family. These included 17 LRR domains (residues 88-634) and one C-terminal LRR domain (LRR-CT, residues 694-745) in the extracellular region, and a TIR domain (residues 801-944) in the cytoplasmic region. Comparison with homologous proteins showed that the deduced CiTLR22 has the highest sequence identity to common carp TLR22 (82.9%). Genomic DNA of CiTLR22 was obtained by long-distance (Ld) PCR and structure analysis revealed that the CiTLR22 gene is encoded by uninterrupted exons. Reverse transcriptase-PCR (RT-PCR) revealed that CiTLR22 is a non-maternal gene. It is prominently expressed in immune relevant tissues such as spleen and head kidney. Quantitative RT-PCR analysis showed that CiTLR22 transcripts were upregulated significantly in immune relevant tissues and blood following grass carp reovirus (GCRV) infection. In the whole genomic sequence, nine single nucleotide polymorphisms (SNPs) were detected. Seven of them were sited in the coding region, and the other two located in the 5' and 3' untranslated region (UTR) respectively. None of the SNPs was associated with the resistance of grass carp to GCRV. These results suggested a role for CiTLR22 in mediating immune protection against viral infection in grass carp.

Gastric Metastasis of Primary Lung Cancer: Case Report and Systematic Review With Pooled Analysis.

Background: Gastric metastasis from lung cancer (GMLC) is a rare occurrence. The clinicopathological characteristics, outcomes, and prognostic factors remain largely elusive. Methods: We conducted a systematic review on case reports and case series of GMLC by scanning MEDLINE, Embase, and ISI Web of Knowledge. Data involving the clinicopathological features, treatment, and outcomes were extracted and analyzed. Survival analysis was performed using Kaplan-Meier method. The Cox proportional hazards regression model was used to identify potential prognostic factors associated with survival. Furthermore, a case of metastatic gastric adenocarcinoma of pulmonary origin with epidermal growth factor receptor (EGFR) L858R+T790M mutation was also described and included. Results: Seventy-eight records involving 114 cases (including ours) were finally included. The median age on admission was 65 years with a male predominance of 79.8%. Lung adenocarcinoma (42.1%), located in the right upper lobe (30.3%), was the most frequent primary tumor. Bleeding (36.7%) and abdominal pain (35.8%) were the two most common symptoms. Endoscopically, gastric lesions were typically presented as elevated lesions with or without volcano-like ulceration, or ulcerative lesions, mostly involving the gastric corpus. The median overall survival time and survival time after diagnosis of metastatic cancer were 11 months [95% confidence interval (CI): 7-14] and 4.5 months (95% CI: 3-9), respectively. The survival analyses revealed that surgical interventions (including lung surgery and/or abdominal surgery) and systemic therapy (including chemotherapy, radiotherapy, and/or targeted therapy) seemed to be positive prognostic factors for both overall survival and survival after diagnosis of metastatic cancer. Conclusions: Clinicians should be alerted to the occurrence of gastric metastasis in lung cancer patients. Comprehensive evaluation and appropriate treatment for specific patients may improve the survival rate of GMLC patients.

Screening and Verification of Molecular Markers and Genes Related to Salt-Alkali Tolerance in Portunus trituberculatus.

Salt-alkali tolerance is one of the important breeding traits of Portunus trituberculatus. Identification of molecular markers linked to salt-alkali tolerance is prerequisite to develop such molecular marker-assisted breeding. In this study, Bulked Segregant Analysis (BSA) was used to screen molecular markers associated with salt-alkali tolerance trait in P. trituberculatus. Two DNA mixing pools with significant difference in salt-alkali tolerance were prepared and 94.83G of high-quality sequencing data was obtained. 855 SNPs and 1051 Indels were firstly selected as candidate markers by BSA analysis, out of which, 20 markers were further selected via △index value (close to 0 or 1) and eight of those were successfully verified. In addition, based on the located information of the markers in genome, eight candidate genes related to salt-alkali tolerance were anchored including ubiquitin-conjugating enzyme, aspartate-tRNA ligase, vesicle-trafficking protein, and so on. qPCR results showed that the expression patterns of all these genes changed significantly after salt-alkali stress, suggesting that they play certain roles in salt-alkali adaptation. Our results will provide applicable markers for molecular marker-assisted breeding and help to clarify the mechanisms of salt-alkali adaptation of P. trituberculatus.

Pan-cancer analysis of ARID family members as novel biomarkers for immune checkpoint inhibitor therapy.

Although immune checkpoint inhibitors (ICIs) have greatly improved cancer treatment, the accuracy of predictive biomarkers for ICI outcomes, such as PD-L1, TMB (tumor mutation burden) or MMR (mismatch repair) deficiency, have not been satisfactory. ARID family members are essential for maintaining the basic process of genomic stability and may serve as novel biomarkers for ICI therapy. A total of 1660 cancer patients who received ICI therapy were included in this pan-cancer analysis. The basic information and TMB values of each patient were collected. Survival analysis based on the Kaplan-Meier (KM) method was performed to explore the relationships between mutations in ARID family members and prognosis in pan-cancer as well as cancer subtypes. Genetic alterations in ARID1A (12%), ARID1B (5%), ARID2 (6%) and ARID5B (2.6%) were identified in multiple cancer types. Patients harboring mutated ARID family members benefited more from ICI therapy (P = .0003). Mutated ARID1A (P = .01), ARID1B (P = .0097) and ARID2 (P = .0054) all serve as compelling biomarkers in predicting the prognosis of ICI treatment. In addition, members of the ARID family were found to be strongly related to the abundance of CD4 + T cells and CD8 + T cells, the expression of PD-L1 and the TMB value in various cancers. Specifically, members of the ARID family could serve as novel biomarkers in multiple malignancies, especially gastrointestinal cancers. ARID family members serve as novel biomarkers for ICI therapy in malignancies. Testing the genomic status of ARID family members could help identify the definite subpopulation that benefits most from ICI treatment.Abbreviations: AT-rich interactive domain (ARID)Switch/sucrose nonfermenting (SWI/SNF)Non-small cell lung cancer (NSCLC)Immune checkpoint inhibitors (ICIs)Tumor microenvironment (TME)Programmed death-ligand 1 (PD-L1)Tumor mutational burden (TMB).

Improved genome assembly of Chinese shrimp (Fenneropenaeus chinensis) suggests adaptation to the environment during evolution and domestication.

A high-quality reference genome is necessary to determine the molecular mechanisms underlying important biological phenomena; therefore, in the present study, a chromosome-level genome assembly of the Chinese shrimp Fenneropenaeus chinensis was performed. Muscle of a male shrimp was sequenced using PacBio platform, and assembled by Hi-C technology. The assembled F. chinensis genome was 1.47 Gb with contig N50 of 472.84 Kb, including 57.73% repetitive sequences, and was anchored to 43 pseudochromosomes, with scaffold N50 of 36.87 Mb. In total, 25,026 protein-coding genes were predicted. The genome size of F. chinensis showed significant contraction in comparison with that of other penaeid species, which is likely related to migration observed in this species. However, the F. chinensis genome included several expanded gene families related to cellular processes and metabolic processes, and the contracted gene families were associated with virus infection process. The findings signify the adaptation of F. chinensis to the selection pressure of migration and cold environment. Furthermore, the selection signature analysis identified genes associated with metabolism, phototransduction, and nervous system in cultured shrimps when compared with wild population, indicating targeted, artificial selection of growth, vision, and behavior during domestication. The construction of the genome of F. chinensis provided valuable information for the further genetic mechanism analysis of important biological processes, and will facilitate the research of genetic changes during evolution.

A chromosome-level genome of Portunus trituberculatus provides insights into its evolution, salinity adaptation and sex determination.

Portunus trituberculatus (Crustacea: Decapoda: Brachyura), commonly known as the swimming crab, is of major ecological importance, as well as being important to the fisheries industry. P. trituberculatus is also an important farmed species in China due to its rapid growth rate and high economic value. Here, we report the genome sequence of the swimming crab, which was assembled at the chromosome scale, covering ~1.2 Gb, with 79.99% of the scaffold sequences assembled into 53 chromosomes. The contig and scaffold N50 values were 108.7 kb and 15.6 Mb, respectively, with 19,981 protein-coding genes. Based on comparative genomic analyses of crabs and shrimps, the C2H2 zinc finger protein family was found to be the only gene family expanded in crab genomes, suggesting it was closely related to the evolution of crabs. The combination of transcriptome and bulked segregant analysis provided insights into the genetic basis of salinity adaptation and rapid growth in P. trituberculatus. In addition, the specific region of the Y chromosome was located for the first time in the genome of P. trituberculatus, and three genes were preliminarily identified as candidate genes for sex determination in this region. Decoding the swimming crab genome not only provides a valuable genomic resource for further biological and evolutionary studies, but is also useful for molecular breeding of swimming crabs.

Differences in and verification of genetic alterations in chemotherapy and immunotherapy for metastatic melanoma.

BACKGROUND: Metastatic melanoma has poor therapeutic response and may present resistance to chemotherapy or immunotherapy. Significant differences are observed in the survival time of patients with metastatic melanoma based on the administration of chemotherapy or immunotherapy; thus, we have explored the important role of specific differential genes between the two therapies in their effect on treatment response in melanoma. METHODS: Metastatic melanoma gene expression data (RNAseq, mutation and methylation) and patient clinical information were downloaded from The Cancer Genome Atlas database and grouped according to chemotherapy or immunotherapy. The differentially expressed genes of the two groups were further screened for signature genes through a protein-protein interaction network and Lasso-Cox regression model. Then, differences in the treatment response, overall survival, mutation and methylation of characteristic genes were compared. Finally, western blot and real-time qPCR technology were used to detect the expression differences of the signature genes in metastatic melanoma tumor tissues in patients undergoing chemotherapy and immunotherapy. RESULTS: The overall survival of the chemotherapy-based treatment group was significantly higher than that of the immunotherapy-based group. The immune infiltration level of immature dendritic cells (DCs) in the chemotherapy group was significantly higher than that in the immunotherapy group. Finally, seven signature genes were selected: CCKBR, KCNJ11, NMU, MMP13, ITGA10, IGFBP1 and CEACAM5. The results of these signature genes were significantly differentiated between the chemotherapy and immunotherapy groups in terms of overall survival and disease progression in response to treatment. In addition, differences in the expression of these genes were verified by western blot and real-time qPCR. CONCLUSION: In this study, significant differences in the expression of signature genes were verified. The findings indicate that immature DCs with potential application value should be considered and high mutation sites of signature genes should be identified to reduce the occurrence of treatment resistance.

Characterization of p53 From the Marine Crab Portunus trituberculatus and Its Functions Under Low Salinity Conditions.

Portunus trituberculatus, or the swimming crab, is tolerant of reduced salinity; however, the molecular mechanism of this tolerance is not clear. Cells can be damaged by hyperosmotic salinity. The protein p53, sometimes referred to as "the guardian of the genome," displays versatile and important functions under changing environmental conditions. Herein, the P. trituberculatus p53 gene (designated as Ptp53) was cloned and studied. The full-length Ptp53 cDNA comprised 1,544bp, with a 1,314bp open reading frame, which encodes a putative polypeptide of 437 amino acids. Quantitative real-time reverse transcription PCR assays revealed ubiquitous expression of Ptp53 in all tissues examined, with the gills showing the highest expression level. Extensive apoptosis was detected under low salinity conditions using terminal deoxynucleotidyl transferase nick-end-labeling staining. Oxidative stress was induced under low salinity conditions, consequently leading to apoptosis. Low salinity stress caused significant upregulation of Ptp53 mRNA and protein levels in the gills. Moreover, compared with that in the control group, the mortality of Ptp53-silenced crabs under low salinity stress was enhanced significantly. Taken together, our findings suggest that Ptp53, via regulation of apoptosis and antioxidant defense, played important functions in the low salinity stress response of the swimming crab.

Full-length transcriptome sequences of ridgetail white prawn Exopalaemon carinicauda provide insight into gene expression dynamics during thermal stress.

Marine heat waves and extreme high temperature become more frequent and intense in these years, which affected the survival of aquaculture animals. The ridgetail white prawn Exopalaemon carinicauda is an important economic species in eastern China, which has remarkable thermal tolerance. However, there has been little study of its thermal-adaptation mechanisms due to the complex genetic structure and unknown genome. To better understand the molecular mechanisms of E. carinicauda to adapt to the changing temperature, a combination of Illumina-based short reads RNA-seq and single molecule real-time-based full-length transcriptome sequencing was used in this study. In total, 17,212 unigenes from high-quality transcripts of E. carinicauda were generated and 14,663 complete ORFs were detected with an average length of 1980 bp. In addition, the transcriptome profiles of E. carinicauda treated with 34 °C heat stress for 6 and 24 h were analyzed. These differentially expressed genes were primarily enriched in oxidation-reduction process (Gene Ontology enrichment, GO) and the pathways of starch and sucrose metabolism (Kyoto Encyclopedia of Genes and Genomes enrichment, KEGG) after 6 h thermal stress, which indicated that E. carinicauda was suffering the attack by reactive oxygen species. After 24 h thermal stress, these differentially expressed genes were enriched in the pathway of lysosome, glycine, serine and threonine metabolism, fatty acid metabolism (KEGG), which indicated the oxidative stress was decreased. Interestingly, 40 genes for hemocyanin were found to be downregulated after 6 h heat stress, which indicated that the immunocompetence of E. carinicauda decreased after short term thermal stress (6 h). After 24 h thermal stress, E. carinicauda showed transcriptional adaptation to high temperature by upregulating of 11 genes encoding molecular chaperones, including HSP40 and HSP90 which were firstly reported to be related to thermal stress in E. carinicauda. These results promote a better understanding of the thermal-adaptation mechanism of E. carinicauda.