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Abstract Endemic to the upper and middle reaches of the Yangtze River in China, elongate loach (Leptobotia elongata) has become a vulnerable species mainly due to overfishing and habitat destruction. Thus far, no genome data of this species are reported. As a result, lacking of such genomic information has restricted practical conservation and utilization of this economic fish. Here, we constructed chromosome-level genome assemblies for both male and female elongate loach by integration of MGI, PacBio HiFi and Hi-C sequencing technologies. Two primary genome assemblies (586-Mb and 589-Mb) were obtained for female and male fishes, respectively. Indeed, 98.22% and 98.61% of the contig sequences were anchored onto 25 chromosomes, with identification of 26.22% and 25.92% repeat contents in both assembled genomes. Meanwhile, a total of 25,215 and 25,253 protein-coding genes were annotated, of which 97.41% and 98.8% could be predicted with functions. Taken together, our genome data presented here provide a valuable genomic resource for in-depth evolutionary and functional research, as well as molecular breeding and conservation of this economic fish species.

Abstract Euglena gracilis (E. gracilis), pivotal in the study of photosynthesis, endosymbiosis, and chloroplast development, is also an industrial microalga for paramylon production. Despite its importance, E. gracilis genome exploration faces challenges due to its intricate nature. In this study, we achieved a chromosome-level de novo assembly (2.37 Gb) using Illumina, PacBio, Bionano, and Hi-C data. The assembly exhibited a contig N50 of 619 Kb and scaffold N50 of 1.12 Mb, indicating superior continuity. Approximately 99.83% of the genome was anchored to 46 chromosomes, revealing structural insights. Repetitive elements constituted 58.84% of the sequences. Functional annotations were assigned to 39,362 proteins, enhancing interpretative power. BUSCO analysis confirmed assembly completeness at 80.39%. This first high-quality E. gracilis genome offers insights for genetics and genomics studies, overcoming previous limitations. The impact extends to academic and industrial research, providing a foundational resource.

Stimulator of interferon genes (STING) has recently been found to play a crucial role in cardiac sterile inflammation and dysfunction. The role of stimulator of interferon genes (STING) in cardiac sterile inflammation and dysfunction has been recently discovered. This study aims to examine the involvement of STING in pathological cardiac remodeling and the mechanisms that govern the activation of the STING pathway. To investigate this, transverse aortic constriction (TAC) was performed on STING knockout mice to induce pressure overload-induced cardiac remodeling. Subsequently, cardiac function, remodeling, and inflammation levels were evaluated. The STING pathway was found to be activated in the pressure overload-stressed heart and angiotensin II (Ang II)-stimulated cardiac fibroblasts. Loss of STING expression led to a significant reduction in inflammatory responses, mitochondrial fragmentation, and oxidative stress in the heart, resulting in attenuated cardiac remodeling and dysfunction. Furthermore, the exacerbation of pressure overload-induced STING-mediated inflammation and pathological cardiac remodeling was observed when mitophagy was suppressed through the silencing of Parkin, an E3 ubiquitin ligase. Taken together, these findings indicate that STING represents a newly identified and significant molecule implicated in the process of pathological cardiac remodeling and that mitophagy is an upstream mechanism that regulates STING activation. Targeting STING may therefore provide a novel therapeutic strategy for pathological cardiac remodeling and heart failure.

Introduction: Mudskippers are a large group of amphibious fishes that have developed many morphological and physiological capacities to live on land. Genomics comparisons of chromosome-level genome assemblies of three representative mudskippers, Boleophthalmus pectinirostris (BP), Periophthalmus magnuspinnatus (PM) and P. modestus (PMO), may be able to provide novel insights into the water-to-land evolution and adaptation. Methods: Two chromosome-level genome assemblies for BP and PM were respectively sequenced by an integration of PacBio, Nanopore and Hi-C sequencing. A series of standard assembly and annotation pipelines were subsequently performed for both mudskippers. We also re-annotated the PMO genome, downloaded from NCBI, to obtain a redundancy-reduced annotation. Three-way comparative analyses of the three mudskipper genomes in a large scale were carried out to discover detailed genomic differences, such as different gene sizes, and potential chromosomal fission and fusion events. Comparisons of several representative gene families among the three amphibious mudskippers and some other teleosts were also performed to find some molecular clues for terrestrial adaptation. Results: We obtained two high-quality haplotype genome assemblies with 23 and 25 chromosomes for BP and PM respectively. We also found two specific chromosome fission events in PM. Ancestor chromosome analysis has discovered a common fusion event in mudskipper ancestor. This fusion was then retained in all the three mudskipper species. A loss of some SCPP (secretory calcium-binding phosphoprotein) genes were identified in the three mudskipper genomes, which could lead to reduction of scales for a part-time terrestrial residence. The loss of aanat1a gene, encoding an important enzyme (arylalkylamine N-acetyltransferase 1a, AANAT1a) for dopamine metabolism and melatonin biosynthesis, was confirmed in PM but not in PMO (as previously reported existence in BP), suggesting a better air vision of PM than both PMO and BP. Such a tiny variation within the genus Periophthalmus exemplifies to prove a step-by-step evolution for the mudskippers’ water-to-land adaptation. Conclusion: These high-quality mudskipper genome assemblies will become valuable genetic resources for in-depth discovery of genomic evolution for the terrestrial adaptation of amphibious fishes.

Abstract Background Immune‐checkpoint inhibitors are now used more commonly in combination than monotherapy as the first‐line choice in patients with unresectable advanced melanoma. Nevertheless, for cases that progressed after the initial combination therapy, the subsequent regimen option can be very difficult. Herein, we reported the efficacy and safety of a triple combination regimen in Chinese unresectable advanced melanoma patients who had poor responses to the first‐line immune therapy. Methods We reviewed the clinical profiles of patients diagnosed with stage IIIC‐IV melanoma between June 1, 2020, and September 30, 2023. The patients who failed the prior immune therapies and received anti‐PD‐1 mono antibody plus interferon(IFN)‐alpha 1b and anlotinib hydrochloride as the second‐line therapy were enrolled in the retrospective analysis. Additionally, we examined the exhaustion of T‐cells using mIHC staining in available tumor samples. Results Fifty‐five patients were included in this study. The median follow‐up period was 13.6 months. The objective response rate evaluated by the investigators was 9.1%(1CR, 4PR). The disease control rate was 47.3%. The median overall survival was 17.6 months, and the median progression‐free survival was 2.8 months. The adverse events rate of any grade was 100%. Grade 3 or 4 irAEs were observed in 29.1% of cases. Multiplex immunohistochemical staining revealed an increased trend of TIM3 expression on tumor‐infiltrating T cells in patients without objective response. Conclusion PD‐1 monoclonal antibody plus interferon‐alpha 1b plus anlotinib showed acceptable tolerability and anticancer benefits in Chinese metastatic melanoma patients as a second‐line therapy.

The yellow catfish (Pelteobagrus fulvidraco) is an economically important species in China and East Asia. However, with the development of high-density farming, bacterial diseases of yellow catfish are becoming increasingly serious. In mammals, autophagy participates in an intracellular innate defense pathway in response to bacterial and viral infections. Among these signaling pathways, stimulator of interferon genes (STING) combines with a ring diode produced by the cyclic GMP-AMP synthase (cGAS) and activates the immune response and autophagy of pathogenic microorganisms. The cGAS-STING signaling pathway in fishes remains largely unexplored. This study found that cGAS and STING exhibited both conserved gene structure and gene synteny in various species of teleosts and diverse species of vertebrates. Expression profiling revealed that the transcription levels of cGAS and STING in immune tissues were significantly up-regulated within 48 h after stimulation with Aeromonas hydrophila or Edwardsiella tarda. Our results demonstrated that the expression levels of TBK1 and IRF3 in the cGAS-STING pathway were significantly up-regulated in the liver, kidney, and spleen, suggesting that this pathway was triggered and activated within 48 h by stimulation with exogenous A. hydrophila. Analyses conducted through Transmission Electron Microscopy (TEM) and marker genes showed that autophagic hyperactivation occurred in the liver, spleen, and kidney of hybrid yellow catfish after infection with A. hydrophila or E. tarda. Taken together, the results indicate that cGAS and STING function are critical mediators of antibacterial immunity in the yellow catfish, and that the cGAS-STING signaling pathway is involved in mediating antibacterial defenses.

Fatty acid desaturases (FADs) are rate-limiting enzymes for the biosynthesis of highly unsaturated fatty acids (HUFAs). As a new member of the FAD family, Fad6 and its roles remain unclear in various teleost fishes. In this study, we identified a fad6 gene from yellow catfish (ycfad6) and determined its spatiotemporal expression patterns and responses to different initial diets and nutritional status in yellow catfish. Our results showed that the open reading frame (ORF) of ycfad6 was 1,080 bp in length, encoding a protein of 359 amino acids. Multiple protein sequences alignment proved that fad6 is highly conserved among diverse vertebrates. Meanwhile, phylogenetic analysis revealed that Southern catfish and yellow catfish were clustered into one branch, supporting evolutionary consistence between the fad6 gene and fish morphology. Moreover, comparisons of genomic synteny and gene structure revealed functional and evolutionary conservation of the fad6 gene in various teleost fishes. Tissue distribution analysis by quantitative RT-PCR demonstrated that the ycfad6 gene was extensively expressed in examined tissues, with higher transcription levels in the heart and liver. Meanwhile, ycfad6 gene was widely expressed in various developmental stages, indicating Fad6 may play important roles in HUFA biosynthesis at early developmental stages in yellow catfish. Functional experiments verified that the transcription of ycfad6 decreased significantly with the extension of feeding time (with egg yolk or Artemia nauplii) at the early developmental stages, indicating that a diet rich in HUFA can remarkably inhibit the transcription of ycfad6 in yellow catfish. In addition, ycfad6 transcription was significantly reduced after a short-term (24-h) or long-term food deprivation (1-week) and then continued to decrease during refeeding, suggesting that nutritional states can affect the transcription of ycfad6, which further regulates the metabolism of HUFAs. Anyway, these fundamental findings provide basic references for further investigating evolutionary and physiological functions of the fad6 gene in yellow catfish as well as in other teleost fishes.

Mitotic kinesin-like protein 2 (MKLP2/KIF20A) is a key mitotic regulator frequently overexpressed in human malignancies and its abundance is positively correlated with poor outcomes of the disease. Despite extensive research on MKLP2 as a potential target for oncology, the development of small-molecule inhibitors specific to MKLP2 remains limited. We have previously identified a benzoazipinone compound, HJ81 as a potent disruptor of Aurora kinase B (AURKB) localization during late mitosis. This study reveals that such disruption results from a failure of AURKB relocation at the onset of anaphase and this phenomenon can be specifically attributed to the disablement of MKLP2, a recognized facilitator of the relocation process. Further optimization of HJ81 leads to identifying compounds such as 12a as promising lead inhibitors of MKLP2-mediated processes, with improved pharmacokinetic properties. 12a inhibits the microtubule-stimulated ATPase activity of the recombinant MKLP2 in vitro. Significant suppression of tumor growth was observed in mice bearing the Calu-6 lung cancer cell line when treated with 12a at a well-tolerated dose. Overall, our findings suggest that benzoazipinone derivatives represent a novel chemical scaffold with the potential to be developed to mimic MKLP2 inhibition for cancer treatment.

Abstract Background Tumor cells frequently suffer from endoplasmic reticulum (ER) stress. Previous studies have extensively elucidated the role of tumorous unfolded protein response in melanoma cells, whereas the effect on tumor immunology and the underlying mechanism remain elusive. Methods Bioinformatics, biochemical assays and pre-clinical mice model were employed to demonstrate the role of tumorous inositol-requiring transmembrane kinase/endoribonuclease 1α (IRE1α) in anti-tumor immunity and the underlying mechanism. Results We firstly found that IRE1α signaling activation was positively associated with the feature of tumor-infiltrating lymphocytes. Then, pharmacological ER stress induction by HA15 exerted prominent anti-tumor effect in immunocompetent mice and was highly dependent on CD8+T cells, paralleled with the reshape of immune cells in tumor microenvironment via tumorous IRE1α-XBP1 signal. Subsequently, tumorous IRE1α facilitated the expression and secretion of multiple chemokines and cytokines via XBP1-NF-κB axis, leading to increased infiltration and anti-tumor capacity of CD8+T cells. Ultimately, pharmacological induction of tumorous ER stress by HA15 brought potentiated therapeutic effect along with anti-PD-1 antibody on melanoma in vivo. Conclusions Tumorous IRE1α facilitates CD8+T cells-dependent anti-tumor immunity and improves immunotherapy efficacy by regulating chemokines and cytokines via XBP1-NF-κB axis. The combination of ER stress inducer and anti-PD-1 antibody could be promising for increasing the efficacy of melanoma immunotherapy.

Abstract Deep-sea hydrothermal vents are usually considered as extreme environments with high pressure, high temperature, scarce food, and chemical toxicity, while many local inhabitants have evolved special adaptive mechanisms for residence in this representative ecosystem. In this study, we constructed a high-quality genome assembly for a novel deep-sea anemone species (Actinostola sp.) that was resident at a depth of 2,971 m in an Edmond vent along the central Indian Ocean ridge, with a total size of 424.3 Mb and a scaffold N50 of 383 kb. The assembled genome contained 265 Mb of repetitive sequences and 20,812 protein-coding genes. Taken together, our reference genome provides a valuable genetic resource for exploring the evolution and adaptive clues of this deep-sea anemone.

Aquaculture, as a rapidly growing industry, plays a significant role in global food production and economic development. However, various infectious diseases, particularly those caused by the gram-negative bacterium Aeromonas hydrophila, pose a critical threat to the sustainability and productivity of world-wide aquaculture operations. Investigating the molecular mechanisms of A. hydrophila infection can help to identify key targets for therapeutic interventions and vaccines, thereby reducing economic losses associated with disease outbreaks. Our present study focuses on wide-bodied sand loach (WBSL, Sinibotia reevesae), a captive-bred commercial fish, to explore detailed molecular effects of exogenous A. hydrophila infection. Firstly, a high-quality chromosome-level haplotype genome with a total length of 774 Mb and 49 chromosomes was assembled, providing valuable insights into its genetic components. Secondly, we isolated and characterized a newfound A. hydrophila F4S strain, examined pathological symptoms of WBSL after infection, and then investigated dynamic gene expressions. Our results revealed that the bacterial infection decreased metabolic activity, causing disruptions in cardiac function, cellular homeostasis, tissue damages, and impaired energy utilization. The immune system responds multifaceted, potentially impacting overall health and survival. In summary, our findings contribute to the understanding of the complicated interactions between WBSL and exogenous A. hydrophila, especially shed lights on the genetic complexity, immune responses, and potential impacts on fish health and survival. Furthermore, this study provides new insights into the mechanisms underlying A. hydrophila infection, and offers potential strategies to offload corresponding pathological effects, not only in WBSL but also in other aquaculture species.

Asia arowana (Scleropages formosus) is an ornamental fish with high economic value, while its sex determination mechanism is still poorly understood. By far, no morphological evidence or molecular marker has been developed for effective distinguishment of genders, which poses a critical challenge to our captive breeding efforts. In this study, we sequenced gonadal transcriptomes of adult Asian arowanas and revealed differential expression profiling of sex-related genes. Based on the comparative transcriptomics analysis of testes (n = 3) and ovaries (n = 3), we identified a total of 8,872 differentially expressed genes (DEGs) and 18,490 differentially expressed transposable elements (TEs) between male and female individuals. Interestingly, the expression of TEs usually has been more significantly testis-biased than related coding genes. As expected, several genes related to females (such as foxl2 and cyp19a1a) are significantly transcribed in the ovary, and some genes related to male gonad development (such as dmrt1, gsdf and amh) are highly expressed in the testis. This sexual dimorphism is valuable for ascertaining the differential expression patterns of sex-related genes and enriching the genetic resources of this economically important species. These valuable genetic materials thereby provide instructive references for gender identification and one-to-one breeding practices so as to expand fish numbers for a rapid elevation of economic value.

Orange spotted grouper (Epinephelus coioides) is an important mariculture fish, and genomic breeding of this grouper species has been hindered due to lack of efficient genotyping tools. Here, we developed a single nucleotide polymorphism (SNP) genotyping technology based on multiplex PCR enrichment capture sequencing, which mainly aims at target area for high-throughput sequencing, and 741 SNPs were designed for genomic selection (GS) of growth and ammonia tolerance traits at the same time. The multiplex PCR enrichment capture sequencing assay showed that the genotyping efficiency was more than 99% in the orange-spotted grouper and the predictive accuracy of body weight and ammonia tolerance traits was 82% and 96%, respectively. More importantly, the average identity of the sequences with these SNPs aligned to the genomes of giant grouper (E. lanceolatus) and brown-marbled grouper (E. fuscoguttatus) were both over 96%. Test data showed that the SNP genotyping efficiency was more than 94% in both giant grouper and brown-marbled grouper. In summary, these results indicated that the development of SNP loci and genotyping approach based on the multiple PCR enrichment capture sequencing are suitable for GS of growth and ammonia tolerance traits in various grouper species, and it would provide technical support for practical grouper breeding.

Abstract The green microalga Haematococcus pluvialis can synthesize high amounts of astaxanthin, which is a valuable antioxidant that has been utilized in human health, cosmetics, and aquaculture. To illustrate detailed molecular clues to astaxanthin yield, we performed PacBio HIFI along with Hi-C sequencing to construct an improved chromosome-level haplotypic genome assembly with 32 chromosomes and a genome size of 316.0 Mb. Its scaffold N50 (942.6 kb) and contig N50 (304.8 kb) have been upgraded remarkably from our previous genome draft, and a total of 32,416 protein-coding genes were predicted. We also established a high-evidence phylogenetic tree from seven representative algae species, with the main aim to calculate their divergence times and identify expanded/contracted gene families. We also characterized genome-wide localizations on chromosomes of some important genes such as five BKTs (encoding beta-carotene ketolases) that are putatively involved in astaxanthin production. In summary, we reported the first chromosome-scale map of H. pluvialis, which provides a valuable genetic resource for in-depth biomedical investigations on this momentous green alga and commercial astaxanthin bioproduction.

Abstract Due to potentially hostile behaviors and elusive habitats, moray eels (Muraenidae) as one group of apex predators in coral reefs all across the globe have not been well investigated. Here, we constructed a chromosome-level genome assembly for the representative Reeve’s moray eel (Gymnothorax reevesii). This haplotype genome assembly is 2.17 Gb in length, and 97.87% of the sequences are anchored into 21 chromosomes. It contains 56.34% repetitive sequences and 23,812 protein-coding genes, of which 96.77% are functionally annotated. This sequenced marine species in Anguilliformes makes a good complement to the genetic resource of eel genomes. It not only provides a genetic resource for in-depth studies of the Reeve’s moray eel, but also enables deep-going genomic comparisons among various eels.

Abstract Melanoma is the most lethal type of skin cancer, originating from the malignant transformation of melanocyte. While the development of targeted therapy and immunotherapy has gained revolutionary advances in potentiating the therapeutic effect, the prognosis of patients with melanoma is still suboptimal. During tumor progression, melanoma frequently encounters stress from both endogenous and exogenous sources in tumor microenvironment. SIRT7 is a nuclear-localized deacetylase of which the activity is highly dependent on intracellular nicotinamide adenine dinucleotide (NAD+), with versatile biological functions in maintaining cell homeostasis. Nevertheless, whether SIRT7 regulates tumor cell biology and tumor immunology in melanoma under stressful tumor microenvironment remains elusive. Herein, we reported that SIRT7 orchestrates melanoma progression by simultaneously promoting tumor cell survival and immune evasion via the activation of unfolded protein response. We first identified that SIRT7 expression was the most significantly increased one in sirtuins family upon stress. Then, we proved that the deficiency of SIRT7 potentiated tumor cell death under stress in vitro and suppressed melanoma growth in vivo. Mechanistically, SIRT7 selectively activated the IRE1α-XBP1 axis to potentiate the pro-survival ERK signal pathway and the secretion of tumor-promoting cytokines. SIRT7 directly de-acetylated SMAD4 to antagonize the TGF-β-SMAD4 signal, which relieved the transcriptional repression on IRE1α and induced the activation of the IRE1α-XBP1 axis. Moreover, SIRT7 up-regulation eradicated anti-tumor immunity by promoting PD-L1 expression via the IRE1α-XBP1 axis. Additionally, the synergized therapeutic effect of SIRT7 suppression and anti-PD-1 immune checkpoint blockade was also investigated. Taken together, SIRT7 can be employed as a promising target to restrain tumor growth and increase the effect of melanoma immunotherapy.

Elongase of very long-chain fatty acid 8 (Elovl8) is a new member identified in the Elovl family that is involved in the synthesis of highly unsaturated fatty acids (HUFAs). However, the evolutionary and physiological roles of this enzyme are still largely unknown. In the present study, the elovl8 gene was identified and characterized from yellow catfish Pelteobagrus fulvidraco, and then its evolutionary and molecular characteristics as well as transcriptional changes in response to various nutritional status were determined. Results showed that the open reading frame (ORF) of elovl8 was 795 bp in length, encoding a protein of 264 amino acids. Multiple sequences alignment showed that the yellow catfish Elovl8 was highly conserved with other homologs in teleosts, sharing similar structural characteristics (including six conserved transmembrane α-helical domains, four conserved elongase motifs, and three highly conserved cysteine residues). Meanwhile, comparisons of genetic synteny confirmed that the elovl8 gene identified from the yellow catfish was the homolog of elovl8b in other teleosts, and thus, the elovl8a gene was lost in the genome of the yellow catfish. Gene structure analysis revealed that the elovl8b gene contained eight exons and seven introns, which was highly conserved in teleosts, implying the functional conservation among various fish species. Tissue distribution analysis detected by real-time quantitative PCR (RT-qPCR) showed that the elovl8 gene was extensively expressed in all detected tissues except eyes, with high expression levels in the intestine and liver. Temporal expression analysis revealed that the expression level of elovl8 was stably expressed in the early 12 h after fertilization, and then dramatically decreased at 24, 48, 72, and 96 h after fertilization, implying that elovl8 is required for HUFA biosynthesis in the early development stages. Functional experiments showed that the expression of the elovl8 gene was stimulated after feeding with egg yolk but was not obviously affected after feeding with halogenated worms, indicating that diets full of HUFAs can inhibit the expression of elovl8 in yellow catfish. Our findings will help us to better understand the evolutionary and functional characteristics of elovl8 in teleosts, and lay a solid basis for investigating the regulation mechanism of HUFA biosynthesis.

【Objective】In this study, we report the adaptability of two representative amphibian species, blue-spotted mudskipper (Boleophthalmus Periophthalmodon, BP) and giant-fin mudskipper (Periophthalmus magnuspinnatus, PM) to live on land, with a view to revealing the key role of gastrointestinal microbiota in the adaptation and specific immunity of mudskippers to amphibious life.【Method】By using 16S amplicon sequencing and metagenomic tec hniques, the microbial composition, diversity, abundance and function of the intestines of amphibian and aquatic fish species were compared. The metagenomic data were utilized to compare two representative mudskippers (BP, PM) and three typical aquatic fish species including Ctenopharyngodon idella (CI), Hypophthalmichthys molitri (HM) and Aristichthys nobilis (AN) and the potential terrestrial marker gastrointestinal microbiota in mudskippers were investigated. In addition, the representative data of gastrointestinal microbiota from marine fish, freshwater fish, amphibians and terrestrial animals were obtained through extensive literature to compare the composition and function of intestinal microbiota in mudskippers.【Result】Firmicutes, Proteobacteria, Bacteroidetes and Fusobacteria were dominant in the gastrointestinal microbiota of mudskippers. The contents of various dominant phyla were strikingly different among BP, PM and aquatic fishes. Th e most significant difference in the gastrointestinal microbiota between the two groups was the proportion of CKC4, a gastrointestinal bacterial phylum that is speculated to be related to host lipid metabolism, which ranged from 4% to 27% in CI, HM, AN but was not found in the mudskipper. In addition, the gastrointestinal microbiota of mudskipper containeds typical bacterial families in terrestrial animals, freshwater and seawater fishes fish and amphibians, which was consistent with their life characteristics at the salt-water interface between water and land. It was also observed that fishes had a higher proportion of Clostridium and Proteobacteria than terrestrial animals, which had more Bacteroides species. More interestingly, certain bacteria strains like S24-7, previously thought to be specific in terrestrial animals, were also identified in both BP and PM.【Conclusion】The results suggest that the gastrointestinal microbial communities of mudskipper are more complex and diverse than those of aquatic fishes, which subsequently stimulate the host to form innate immune receptor gene families with higher diversity and more copy numbers through pathogen-related molecular pattern (PAMP) to a more complicated amphibian environment.

Progranulin (PGRN) is a growth factor that is involved in the progression of multiple tumors. However, the effects and molecular mechanisms by which PGRN induces lung cancer remain unclear. The expression level of PGRN was analyzed by conducting immunohistochemistry of the histological sections of lung tissues from non-small-cell lung carcinoma (NSCLC) patients. The proliferation, apoptosis, migration, and invasion of NSCLC cells were assessed by the MTT assay, Western blot, degree of wound healing, and Transwell assays. A nude mouse xenograft model was used to validate the role of PGRN in vivo. The expression level of PGRN was higher in male patients with lung adenocarcinoma than in those with lung squamous cell carcinoma; by contrast, no difference was observed in female patients. The overexpression of PGRN promoted the proliferation and anti-apoptosis of H520 (derived from lung squamous cell carcinoma) cells, whereas knockdown of PGRN inhibited the proliferation and anti-apoptosis of A549 (derived from lung adenocarcinoma) cells. Copanlisib (targeting PI3K) inhibited the increase in the expression of cell anti-apoptosis marker Bcl-2 induced by rhPGRN protein; the PI3K agonist 740 Y–P partially reversed the decrease in Bcl-2 expression induced by PGRN deficiency in both A549 and H520 cells. PGRN increased the expression of Ki-67, PCNA, and Bcl-2 in vivo. PGRN inhibited cell apoptosis depending on the PI3K/Akt/Bcl-2 signaling axis; PGRN positivity correlated with lung adenocarcinoma. PGRN is a potential biomarker for the treatment and diagnosis of NSCLC, especially in lung adenocarcinoma.

Background: S100 calcium-binding protein A9 (S100A9) is a danger-associated molecular pattern molecule that mediates the inflammatory response. Inflammation is essential in aging-related cardiovascular diseases. However, less is known regarding the role of S100A9 in vascular aging. Methods: S100A9 null mice were used to investigate the role of S100A9 in aging-related pathologies. Artery rings were used to measure the functional characteristics of vascular with a pressurized myograph. Telomere length, Sirtuin activity, oxidative stress, and endothelial nitric oxide synthetase (eNOS) activity were used to elevate vascular senescence. Intraperitoneal glucose tolerance (IPGTT) and insulin sensitivity test (IST) were employed to investigate the effects of S100A9 on insulin resistance. Inflammation response was reflected by the concentration of inflammatory cytokines. The Toll-like receptor 4 (TLR4) and receptor for advanced glycation end products (RAGE) inhibitors were used to identify the downstream molecular mechanisms of S100A9 in aging-induced senescence in endothelial cells. Results: S100A9 expression in vascular increased with aging in mice and humans. Deficiency of S100A9 alleviated vascular senescence in aged mice, as evidenced by increased telomere length, Sirtuin activity, and eNOS activity. Meanwhile, S100A9 knockout improved endothelium-dependent vasodilatation and endothelial continuity in aged mice. Moreover, the increased insulin resistance, oxidative stress, and inflammation were mitigated by S100A9 deletion in aged mice. In vitro, S100A9 induced senescence in endothelial cells, and that effect was blunted by TLR4 but not RAGE inhibitors. Conclusion: The present study suggested that S100A9 may contribute to aging-related pathologies and endothelial dysfunction via the TLR4 pathway. Therefore, targeting S100A9/TLR4 signaling pathway may represent a crucial therapeutic strategy to prevent age-related cardiovascular diseases.

Background: Largemouth bass (Micropterus Salmoides) is an economically important fish species in China. Most research has focused on its growth, disease resistance, and nutrition improvement. However, the sex-determining genes in largemouth bass are still unclear. The transforming growth factor-beta (TGF-β) gene family, including amh, amhr2 and gsdf, plays an important role in the sex determination and differentiation of various fishes. These genes are potentially involved in sex determination in largemouth bass. Methods: We performed a systematic analysis of 5 sex-related genes (amh, amhr2, gsdf, cyp19a1, foxl2) in largemouth bass using sequence alignment, collinearity analysis, transcriptome, and quantitative real-time polymerase chain reaction (qRT-PCR). This included a detailed assessment of their sequences, gene structures, evolutionary traits, and gene transcription patterns in various tissues including gonads, and at different developmental stages. Results: Comparative genomics revealed that the 5 sex-related genes were highly conserved in various fish genomes. These genes did not replicate, mutate or lose in largemouth bass. However, some were duplicated (amh, amhr2 and gsdf), mutated (gsdf) or lost (amhr2) in other fishes. Some genes (e.g., gsdf) showed significant differences in genomic sequence between males and females, which may contribute to sex determination and sex differentiation in these fishes. qRT-PCR was applied to quantify transcription profiling of the 5 genes during gonadal development and in the adult largemouth bass. Interestingly, amh, amhr2 and gsdf were predominantly expressed in the testis, while cyp19a1 and foxl2 were mainly transcribed in the ovary. All 5 sex-related genes were differentially expressed in the testes and ovaries from the 56th day post-fertilization (dpf). We therefore speculate that male/female differentiation in the largemouth bass may begin at this critical time-point. Examination of the transcriptome data also allowed us to screen out several more sex-related candidate genes. Conclusions: Our results provide a valuable genetic resource for investigating the physiological functions of these 5 sex-related genes in sex determination and gonadal differentiation, as well as in the control of gonad stability in adult largemouth bass.

The venoms of various sea anemones are rich in diverse toxins, which usually play a dual role in capturing prey and deterring predators. However, the complex components of such venoms have not been well known yet. Here, venomics of integrating transcriptomic and proteomic technologies was applied for the first time to identify putative protein and peptide toxins from different tissues of the representative sea anemone, Heteractis magnifica. The transcriptomic analysis of H. magnifica identified 728 putative toxin sequences, including 442 and 381 from the tentacles and the column, respectively, and they were assigned to 68 gene superfamilies. The proteomic analysis confirmed 101 protein and peptide toxins in the venom, including 91 in the tentacles and 39 in the column. The integrated venomics also confirmed that some toxins such as the ShK-like peptides and defensins are co-expressed in both the tentacles and the column. Meanwhile, a homology analysis was conducted to predict the three-dimensional structures and potential activity of seven representative toxins. Altogether, this venomics study revealed the venom complexity of H. magnifica, which will help deepen our understanding of cnidarian toxins, thereby supporting the in-depth development of valuable marine drugs.

Marine polychaetes represent an extremely rich and underexplored source of novel families of antimicrobial peptides (AMPs). The rapid development of next generation sequencing technologies and modern bioinformatics approaches allows us to apply them for characterization of AMP-derived genes and the identification of encoded immune-related peptides with the aid of genome and transcriptome mining. Here, we describe a universal bioinformatic approach based on the conserved BRICHOS domain as a search query for the identification of novel structurally unique AMP families in annelids. In this paper, we report the discovery of 13 novel BRICHOS-related peptides, ranging from 18 to 91 amino acid residues in length, in the cosmopolitan marine worm Heteromastus filiformis with the assistance of transcriptome mining. Two characteristic peptides with a low homology in relation to known AMPs—the α-helical amphiphilic linear peptide, consisting of 28 amino acid residues and designated as HfBRI-28, and the 25-mer β-hairpin peptide, specified as HfBRI-25 and having a unique structure stabilized by two disulfide bonds—were obtained and analyzed as potential antimicrobials. Interestingly, both peptides showed the ability to kill bacteria via membrane damage, but mechanisms of their action and spectra of their activity differed significantly. Being non-cytotoxic towards mammalian cells and stable to proteolysis in the blood serum, HfBRI-25 was selected for further in vivo studies in a lethal murine model of the Escherichia coli infection, where the peptide contributed to the 100% survival rate in animals. A high activity against uropathogenic strains of E. coli (UPEC) as well as a strong ability to kill bacteria within biofilms allow us to consider the novel peptide HfBRI-25 as a promising candidate for the clinical therapy of urinary tract infections (UTI) associated with UPEC.

Measurement(s) whole genome sequencing Technology Type(s) Illumina Sequencing • Oxford Nanopore Sequencing

BackgroundThis study aimed to access whether serum human epididymis protein 4 (HE4) level could identify lupus nephritis (LN) pathological classes in adults and children.MethodsThe serum HE4 levels of 190 healthy subjects and 182 patients with systemic lupus erythematosus (SLE) (61 adult-onset LN [aLN], 39 childhood-onset LN [cLN], and 82 SLE without LN) were determined using Architect HE4 kits and an Abbott ARCHITECT i2000SR Immunoassay Analyzer.ResultsSerum HE4 level was significantly higher in the aLN patients (median, 85.5 pmol/L) than in the patients with cLN (44 pmol/L, P < 0.001) or SLE without LN (37 pmol/L, P < 0.001), or the healthy controls (30 pmol/L, P < 0.001). Multivariate analysis showed that serum HE4 level was independently associated with aLN. Stratified by LN class, serum HE4 level was significantly higher in the patients with proliferative LN (PLN) than in those with non-PLN, and this difference was found only in aLN (median, 98.3 versus 49.3 pmol/L, P = 0.021) but not in cLN. Stratified by activity (A) and chronicity (C) indices, the aLN patients with class IV (A/C) possessed significantly higher serum HE4 levels than those with class IV (A) (median, 195.5 versus 60.8 pmol/L, P = 0.006), and this difference was not seen in the class III aLN or cLN patients.ConclusionSerum HE4 level is elevated in patients with class IV (A/C) aLN. The role of HE4 in the pathogenesis of chronic lesions of class IV aLN needs further investigation.

Toll‐like receptor 1 (TLR1) mediates the innate immune response to a variety of microbes through recognizing cell wall components (such as bacterial lipoproteins) in mammals. However, the detailed molecular mechanism of TLR1 involved in pathogen immunity in the representative hybrid yellow catfish (Pelteobagrus fulvidraco ♀ × P. vachelli ♂) has not been well studied. In the present study, we identified the TLR1 gene from the hybrid yellow catfish, and further comparative synteny data from multiple species confirmed that the TLR1 gene is highly conserved in teleosts. Phylogenetic analysis revealed distinguishable TLR1s in diverse taxa, suggesting consistence in evolution of the TLR1 proteins with various species. Structural prediction indicated that the three-dimensional structures of TLR1 proteins are relatively conserved among different taxa. Positive selection analysis showed that purifying selection dominated the evolutionary process of TLR1s and TLR1-TIR domain in both vertebrates and invertebrates. Expression pattern analysis based on the tissue distribution showed that TLR1 mainly transcribed in the gonad, gallbladder and kidney, and the mRNA levels of TLR1 in kidney were remarkably up-regulated after Aeromonas hydrophila stimulation, indicating that TLR1 participates in the inflammatory responses to exogenous pathogen infection in hybrid yellow catfish. Homologous sequence alignment and chromosomal location indicated that the TLR signaling pathway is very conserved in the hybrid yellow catfish. The expression patterns of TLR signaling pathway related genes (TLR1- TLR2 - MyD88 - FADD - Caspase 8) were consistent after pathogen stimulation, revealing that the TLR signaling pathway is triggered and activated after A. hydrophila infection. Our findings will lay a solid foundation for better understanding the immune roles of TLR1 in teleosts, as well as provide basic data for developing strategies to control disease outbreak in hybrid yellow catfish.

Abstract Background Calcific aortic valve disease (CAVD) is the most commonly valvular disease in the western countries initiated by inflammation and abnormal calcium deposition. Currently, there is no clinical drug for CAVD. Neutrophil elastase (NE) plays a causal role in inflammation and participates actively in cardiovascular diseases. However, the effect of NE on valve calcification remains unclear. So we next explore whether it is involved in valve calcification and the molecular mechanisms involved. Methods NE expression and activity in calcific aortic valve stenosis (CAVD) patients (n = 58) and healthy patients (n = 30) were measured by enzyme-linked immunosorbent assay (ELISA), western blot and immunohistochemistry (IHC). Porcine aortic valve interstitial cells (pVICs) were isolated and used in vitro expriments. The effects of NE on pVICs inflammation, apoptosis and calcification were detected by TUNEL assay, MTT assay, reverse transcription polymerase chain reaction (RT-PCR) and western blot. The effects of NE knockdown and NE activity inhibitor Alvelestat on pVICs inflammation, apoptosis and calcification under osteogenic medium induction were also detected by RT-PCR, western blot, alkaline phosphatase staining and alizarin red staining. Changes of Intracellular signaling pathways after NE treatment were measured by western blot. Apolipoprotein E−/− (APOE−/−) mice were employed in this study to establish the important role of Alvelestat in valve calcification. HE was used to detected the thickness of valve. IHC was used to detected the NE and α-SMA expression in APOE−/− mice. Echocardiography was employed to assess the heat function of APOE−/− mice. Results The level and activity of NE were evaluated in patients with CAVD and calcified valve tissues. NE promoted inflammation, apoptosis and phenotype transition in pVICs in the presence or absence of osteogenic medium. Under osteogenic medium induction, NE silencing or NE inhibitor Alvelestat both suppressed the osteogenic differentiation of pVICs. Mechanically, NE played its role in promoting osteogenic differentiation of pVICs by activating the NF-κB and AKT signaling pathway. Alvelestat alleviated valve thickening and decreased the expression of NE and α-SMA in western diet-induced APOE−/− mice. Alvelestat also reduced NE activity and partially improved the heart function of APOE−/−mice. Conclusions Collectively, NE is highly involved in the pathogenesis of valve calcification. Targeting NE such as Alvelestat may be a potential treatment for CAVD.

Abstract Background The explosive radiation and diversification of the advanced snakes (superfamily Colubroidea) was associated with changes in all aspects of the shared venom system. Morphological changes included the partitioning of the mixed ancestral glands into two discrete glands devoted for production of venom or mucous respectively, as well as changes in the location, size and structural elements of the venom-delivering teeth. Evidence also exists for homology among venom gland toxins expressed across the advanced snakes. However, despite the evolutionary novelty of snake venoms, in-depth toxin molecular evolutionary history reconstructions have been mostly limited to those types present in only two front-fanged snake families, Elapidae and Viperidae. To have a broader understanding of toxins shared among extant snakes, here we first sequenced the transcriptomes of eight taxonomically diverse rear-fanged species and four key viperid species and analysed major toxin types shared across the advanced snakes. Results Transcriptomes were constructed for the following families and species: Colubridae - Helicops leopardinus, Heterodon nasicus, Rhabdophis subminiatus; Homalopsidae – Homalopsis buccata; Lamprophiidae - Malpolon monspessulanus, Psammophis schokari, Psammophis subtaeniatus, Rhamphiophis oxyrhynchus; and Viperidae – Bitis atropos, Pseudocerastes urarachnoides, Tropidolaeumus subannulatus, Vipera transcaucasiana. These sequences were combined with those from available databases of other species in order to facilitate a robust reconstruction of the molecular evolutionary history of the key toxin classes present in the venom of the last common ancestor of the advanced snakes, and thus present across the full diversity of colubroid snake venoms. In addition to differential rates of evolution in toxin classes between the snake lineages, these analyses revealed multiple instances of previously unknown instances of structural and functional convergences. Structural convergences included: the evolution of new cysteines to form heteromeric complexes, such as within kunitz peptides (the beta-bungarotoxin trait evolving on at least two occasions) and within SVMP enzymes (the P-IIId trait evolving on at least three occasions); and the C-terminal tail evolving on two separate occasions within the C-type natriuretic peptides, to create structural and functional analogues of the ANP/BNP tailed condition. Also shown was that the de novo evolution of new post-translationally liberated toxin families within the natriuretic peptide gene propeptide region occurred on at least five occasions, with novel functions ranging from induction of hypotension to post-synaptic neurotoxicity. Functional convergences included the following: multiple occasions of SVMP neofunctionalised in procoagulant venoms into activators of the clotting factors prothrombin and Factor X; multiple instances in procoagulant venoms where kunitz peptides were neofunctionalised into inhibitors of the clot destroying enzyme plasmin, thereby prolonging the half-life of the clots formed by the clotting activating enzymatic toxins; and multiple occasions of kunitz peptides neofunctionalised into neurotoxins acting on presynaptic targets, including twice just within Bungarus venoms. Conclusions We found novel convergences in both structural and functional evolution of snake toxins. These results provide a detailed roadmap for future work to elucidate predator–prey evolutionary arms races, ascertain differential clinical pathologies, as well as documenting rich biodiscovery resources for lead compounds in the drug design and discovery pipeline.

The mitotic regulator, Aurora kinase B (AURKB), is frequently overexpressed in malignancy and is a target for therapeutic intervention. The compound, LXY18, is a potent, orally available small molecule that inhibits the proper localization of AURKB during late mitosis, without affecting its kinase activity. In this study, we demonstrate that LXY18 elicits apoptosis in cancer cells derived from various indications, but not in non-transformed cell lines. The apoptosis is p53-independent, triggered by a prolonged mitotic arrest and occurs predominantly in mitosis. Some additional cells succumb post-mitotic slippage. We also demonstrate that cancer cell lines refractory to AURKB kinase inhibitors are sensitive to LXY18. The mitotic proteins MKLP2, NEK6, NEK7 and NEK9 are known regulators of AURKB localization during the onset of anaphase. LXY18 fails to inhibit the catalytic activity of these AURKB localization factors. Overall, our findings suggest a novel activity for LXY18 that produces a prolonged mitotic arrest and lethality in cancer cells, leaving non-transformed cells healthy. This new activity suggests that the compound may be a promising drug candidate for cancer treatment and that it can also be used as a tool compound to further dissect the regulatory network controlling AURKB localization.

We report the first draft genome assembly for Prochilodus magdalenae, the leading representative species of the Prochilodontidae family in Colombia. This 1.2-Gb assembly, with a GC content of 42.0% and a repetitive content of around 31.0%, is in the range of previously reported characid species genomes. Annotation identified 34,725 nuclear genes, and BUSCO completeness value was 94.9%. Gene ontology and primary metabolic pathway annotations indicate similar gene profiles for P. magdalenae and the closest species with annotated genomes: blind cave fish (Astyanax mexicanus) and red piranha (Pygocentrus nattereri). A comparative analysis showed similar genome traits to other characid species. The fully sequenced and annotated mitochondrial genome reproduces the taxonomic classification of P. magdalenae and confirms the low mitochondrial genetic divergence inside the Prochilodus genus. Phylogenomic analysis, using nuclear single-copy orthologous genes, also confirmed the evolutionary position of the species. This genome assembly provides a high-resolution genetic resource for sustainable P. magdalenae management in Colombia and, as the first genome assembly for the Prochilodontidae family, will contribute to fish genomics throughout South America.

Individual cone snail (Conus sp.) contains thousands of bioactive peptides, but there are limited studies on its antimicrobial peptides (AMPs). Here, we investigated AMPs along with AMP-derived genes in the representative Chinese tubular cone snail (C. betulinus) by integration of our previously published multi-omics (genomics, transcriptomics, and peptidomics) data. We identified a total of 466 putative AMP-derived genes from the genome, and most of them were annotated as histones. While at the mRNA level, only 77 AMP-derived genes were confirmed with transcriptomic evidence, among them Ubiquicidin, cgUbiquitin, Ap, and VK10 were the most abundant transcripts in the venom gland. In addition, 30 AMPs were further validated by peptidomics data. After in silico analysis including 3D modeling, 11 putative AMPs were chemically synthesized for an in vitro assessment, and eight peptides were proved with good antifungal activity. In summary, this work systematically characterized the AMP repertoire in C. betulinus. This valuable genetic resource provides a solid foundation for in-depth therapeutic applications and drug development.

Marine cone snail venoms represent a vast library of bioactive peptides with proven potential as research tools, drug leads, and therapeutics. In this study, a transcriptome library of four different organs, namely radular sheath, venom duct, venom gland, and salivary gland, from piscivorous Conus striatus was constructed and sequenced using both Illumina next-generation sequencing (NGS) and PacBio third-generation sequencing (TGS) technologies. A total of 428 conotoxin precursor peptides were retrieved from these transcriptome data, of which 413 conotoxin sequences assigned to 13 gene superfamilies, and 15 conotoxin sequences were classified as unassigned families. It is worth noting that there were significant differences in the diversity of conotoxins identified from the NGS and TGS data: 82 conotoxins were identified from the NGS datasets while 366 conotoxins from the TGS datasets. Interestingly, we found point mutations in the signal peptide sequences of some conotoxins with the same mature sequence. Therefore, TGS broke the traditional view of the conservation of conotoxin signal peptides and the variability of mature peptides obtained by NGS technology. These results shed light on the integrated NGS and TGS technologies to mine diverse conotoxins in Conus species, which will greatly contribute to the discovery of novel conotoxins and the development of new marine drugs.

The economically important Southern bluefin tuna (Thunnus maccoyii) is a world-famous fast-swimming fish, but its genomic information is limited. Here, we performed whole genome sequencing and assembled a draft genome for Southern bluefin tuna, aiming to generate useful genetic data for comparative functional prediction. The final genome assembly is 806.54 Mb, with scaffold and contig N50 values of 3.31 Mb and 67.38 kb, respectively. Genome completeness was evaluated to be 95.8%. The assembled genome contained 23,403 protein-coding genes and 236.1 Mb of repeat sequences (accounting for 29.27% of the entire assembly). Comparative genomics analyses of this fast-swimming tuna revealed that it had more than twice as many hemoglobin genes (18) as other relatively slow-moving fishes (such as seahorse, sunfish, and tongue sole). These hemoglobin genes are mainly localized in two big clusters (termed as “MNˮ and “LAˮ respectively), which is consistent with other reported fishes. However, Thr39 of beta-hemoglobin in the MN cluster, conserved in other fishes, was mutated as cysteine in tunas including the Southern bluefin tuna. Since hemoglobins are reported to transport oxygen efficiently for aerobic respiration, our genomic data suggest that both high copy numbers of hemoglobin genes and an adjusted function of the beta-hemoglobin may support the fast-swimming activity of tunas. In summary, we produced a primary genome assembly and predicted hemoglobin-related roles for the fast-swimming Southern bluefin tuna.

Liver is an important organ for regulating glucose and lipid metabolism. Recent studies have shown that bone morphogenetic proteins (BMPs) may play important roles in regulating glucose and lipid metabolism. In our previous studies, we demonstrated that BMP4 significantly inhibits hepatic steatosis and lowers serum triglycerides, playing a protective role against the progression of non-alcoholic fatty liver disease (NAFLD). However, the direct impact of BMP4 on hepatic glucose metabolism is poorly understood. Here, we investigated the regulatory roles of BMP4 in hepatic glucose metabolism. Through a comprehensive analysis of the 14 types of BMPs, we found that BMP4 was one of the most potent BMPs in promoting hepatic glycogen accumulation, reducing the level of glucose in hepatocytes and effecting the expression of genes related to glucose metabolism. Mechanistically, we demonstrated that BMP4 reduced the hepatic glucose levels through the activation of mTORC2 signaling pathway in vitro and in vivo. Collectively, our findings strongly suggest that BMP4 may play an essential role in regulating hepatic glucose metabolism. This knowledge should aid us to understand the molecular pathogenesis of NAFLD, and may lead to the development of novel therapeutics by exploiting the inhibitory effects of BMPs on hepatic glucose and lipid metabolism.

Abstract Background Global climate oscillation, as a selection dynamic, is an ecologically important element resulting in global biodiversity. During the glacial geological periods, most organisms suffered detrimental selection pressures (such as food shortage and habitat loss) and went through population declines. However, during the mild interglacial periods, many species re-flourished. These temporal dynamics of effective population sizes (N e ) provide essential information for understanding and predicting evolutionary outcomes during historical and ongoing global climate changes. Results Using high-quality genome assemblies and corresponding sequencing data, we applied the Pairwise Sequentially Markovian Coalescent (PSMC) method to quantify N e changes of twelve representative teleost species from approximately 10 million years ago (mya) to 10 thousand years ago (kya). These results revealed multiple rounds of population contraction and expansion in most of the examined teleost species during the Neogene and the Quaternary periods. We observed that 83% (10/12) of the examined teleosts had experienced a drastic decline in N e before the last glacial period (LGP, 110–12 kya), slightly earlier than the reported pattern of N e changes in 38 avian species. In comparison with the peaks, almost all of the examined teleosts maintained long-term lower N e values during the last few million years. This is consistent with increasingly dramatic glaciation during this period. Conclusion In summary, these findings provide a more comprehensive understanding of the historical N e changes in teleosts. Results presented here could lead to the development of appropriate strategies to protect species in light of ongoing global climate changes.

Vibrio parahaemolyticus is an important marine pathogen that causes inflammation and even death in teleost fishes. It has brought significant economic losses to the aquaculture industry as well as high risks to the sustainable development of marine fisheries. In the present study, the fish Tetraodon nigroviridis and the bacterial pathogen Vibrio parahaemolyticus were used to explore the molecular mechanisms underlying the immune response of T. nigroviridis to V. parahaemolyticus exogenous infection. The microRNA (miRNA)–mRNA–protein omics and corresponding experimental validation, followed by comparative analysis, revealed several differentially expressed genes involved in various components of the immune system, including the following: complement system, chemokines, lysosomes, phagocytes, B-cell receptor signaling pathway, T-cell receptor signaling pathway, Janus kinase-signal transducer and activator of transcription (JAK-STAT) signaling pathway, and phospholipid metabolism, among others. Especially, the complements component 3 (C3) gene and protein expression levels were significantly higher after V. parahaemolyticus infection, and miRNAs targeting C3, including mir-6089-y, mir-460-y, and mir-1584-x, were significantly down-regulated. The gene and protein expression levels of complement 1 subunit qA (C1qA) were significantly down-regulated, while mir-203 targeting C1qA was significantly up-regulated. Overall, four complement genes (C1qA, IG, C3, and C5), which are key genes in the classical pathway of complement system activation for inflammatory response, were identified. Evolutionary analysis suggested that T. nigroviridis, acquired an increased ability to recognize pathogens by evolving a more complex complement system than terrestrial vertebrates. In addition, quantitative real-time polymerase chain reaction showed high consistency with the obtained multi-omics results, indicating the reliability of the sequencing data generated in the present study. In summary, our findings can serve as a fundamental basis for further in-depth multi-omics studies on the inflammatory processes of aquatic pathogens hindering fish sustainable production.

Recently, the prevalence of hypertension has become a global challenge. Therefore, tremendous efforts have been made to identify and purify antihypertensive peptides (AHTPs) from food-derived proteins to aid the discovery of new hypotensive drugs with fewer side effects. In this study, we performed high-throughput prediction of AHTPs based on multi-omics data, providing an overview of AHTPs in the lined seahorse proteins and suggesting their potential application as bioactive agents to lower blood pressure. We identified 14,695 AHTP-derived genes in the lined seahorse, and most of them were supported by transcriptomic evidence, whereas only 495 genes were further detected by proteome sequencing. Among these predicted AHTP-derived genes, the longest titin had the most hits with 104 AHTPs, some of which were clustered in exon 158, 194, and 204. Another AHTP-rich group was in the collagen family, and four AHTP-rich collagens exhibited much higher transcription in the pouch than in other examined tissues, including brain, testis, and embryos. Additionally, antihypertensive triplets, comprised of the permutations of Gly, Pro, and Lys, prevailed in all collagen sequences due to the representative XaaYaaGly repeat units. In summary, our present findings provide a solid basis for understanding the abundance of various AHTPs in the lined seahorse as well as shed light on the development of antihypertensive products and drugs using seahorses as an important resource.

Comprising a major clade of Anura, toads produce and secrete numerous toxins from both the parotoid glands behind their eyes and their dorsal skin. These toxins, made of various proteins and compounds, possess pharmacological potential to be repurposed to benefit human health. However, the detailed genetic regulation of toad toxin production is still poorly understood. A recent publication uncovering the genome of the representative Asiatic toad (Bufo gargarizans) provides a good reference to resolve this issue. In the present study, we sequenced the transcriptomes of parotoid gland, dorsal skin and liver from the Asiatic toad. Combining our data with 35 previously published transcriptomes across eight different tissues from the same species but from different locations, we constructed a comprehensive gene co-expression network of the Asiatic toad with the assistance of the reference genome assembly. We identified 2,701 co-expressed genes in the toxin-producing tissues (including parotoid gland and dorsal skin). By comparative genomic analysis, we identified 599 expanded gene families with 2,720 genes. Through overlapping these co-expressed genes in the toad toxin-producing tissues, we observed that three cytochrome P450 (Cyp) family members (Cyp27a1, Cyp2c29, and Cyp2c39) were significantly enriched in pathways related to cholesterol metabolism. Cholesterol is a critical precursor to steroids, and the known main steroidal toxins of bufadienolides are considered as the major bioactive components in the parotoid glands of Asiatic toad. We found 3-hydroxy-methylglutaryl CoA reductase (hmgcr), encoding the major rate-limiting enzyme for cholesterol biosynthesis, appears with multiple copies in both Asiatic toad and common toad, possibly originating from a tandem duplication event. The five copies of hmgcr genes consistently displayed higher transcription levels in the parotoid gland when compared with the abdominal skin, suggesting it as a vital candidate gene in the involvement of toad toxin production. Taken together, our current study uncovers transcriptomic and gene-family dynamic evidence to reveal the vital role of both expanded gene copies and gene expression changes for production of toad toxins.

Midas cichlid (Amphilophus citrinellus), a popular aquarium fish, attracts extensive attention from worldwide biologists mainly due to its morphological polymorphism (dark versus gold). Continuous efforts have therefore been paid to address mechanisms of its coloration variants, while it is far away from the detailed illustration of a clear regulatory network. Some limits may come from the absence of a high-quality genome assembly and a relatively accurate gene set. In this study, we sequenced about 149 Gb of nucleotide sequences of Midas cichlid, generating a genome assembly with a total size of 933.5 Mb, which exhibits a good genome continuity with a contig N50 of 10.5 Mb. A total of 25,911 protein-coding genes were annotated and about 90% completeness was achieved, which helps to build a good gene pool for understanding expressional differences of color variation. With the assistance of the final gene set, we identified a total of 277 differential expressional genes (DEGs), of which 97 up- and 180 downregulated were determined in dark-vs-gold comparisons. Two protein-protein interaction (PPI) networks were constructed from these DEGs, and three key functional modules were classified. Hub genes within each module were evaluated, and we found that the third key module contains tyrp1b, oca2, pmela, tyr, and slc24a5, which were previously proven to be associated with melanin formation. Two downregulated DEGs (myl1 and pgam2) in the first key module may be involved in muscle movement and spermatogenesis, implying that certain side effects could result from the morphological polymorphism. The first key module, consisting of proteins encoded by upregulated DEGs that were associated with MAPK signaling, Toll-like receptor signaling, and gonadotropin-releasing hormone pathways, may contribute to a negative upstream regulation or downstream influence on melanin biosynthesis. Taken together, our new genome assembly and gene annotation of Midas cichlid provide a high-quality genetic resource for biological studies on this species, and the newly identified key networks and hub genes in dark-vs-gold comparisons enhance our understanding of the transcriptional regulatory mechanisms underlying coloration changes not only in Midas cichlid but also in other fishes from freshwater to marine ecosystems.

Abstract Background Despite recent advances in diagnostic and therapeutic approaches for gastric cancer (GC), the survival of patients with advanced GC remains very low. Islet-1 (ISL1) is a LIM-homeodomain transcription factor, which is upregulated and promotes cell proliferation in GC. The exact mechanism by which ISL1 influences GC development is unclear. Methods Co-immunoprecipitation (co-IP) and glutathione S-transferase (GST)-pulldown assays were employed to evaluate the interaction of ISL1 with CDK1. Western blot and immunohistochemistry analyses were performed to evaluate the ability of CDK1 to phosphorylate ISL1 at Ser 269 in GC cell and tissue specimens. Chromatin immunoprecipitation (ChIP), ChIP re-IP, luciferase reporter, and CCK-8 assays were combined with flow cytometry cell cycle analysis to detect the transactivation potency of ISL1-S269-p and its ability to promote cell proliferation. The self-stability and interaction with CDK1 of ISL1-S269-p were also determined. Results ISL1 is phosphorylated by CDK1 at serine 269 (S269) in vivo. Phosphorylation of ISL1 by CDK1 on serine 269 strengthened its binding on the cyclin B1 and cyclin B2 promoters and increased its transcriptional activity in GC. Furthermore, CDK1-dependent phosphorylation of ISL1 correlated positively with ISL1 protein self-stability in NIH3T3 cells. Conclusions ISL1-S269-p increased ISL1 transcriptional activity and self-stability while binding to the cyclinB1 and cyclinB2 promoters promotes cell proliferation. ISL1-S269-p is therefore crucial for tumorigenesis and potentially a direct therapeutic target for GC.

Abstract Background Although almost all extant spider species live in terrestrial environments, a few species live fully submerged in freshwater or seawater. The intertidal spiders (genus Desis) built silk nests within coral crevices can survive submerged in high tides. The diving bell spider, Argyroneta aquatica, resides in a similar dynamic environment but exclusively in freshwater. Given the pivotal role played by mitochondria in supplying most energy for physiological activity via oxidative phosphorylation and the environment, herein we sequenced the complete mitogenome of Desis jiaxiangi to investigate the adaptive evolution of the aquatic spider mitogenomes and the evolution of spiders. Results We assembled a complete mitogenome of the intertidal spider Desis jiaxiangi and performed comparative mitochondrial analyses of data set comprising of Desis jiaxiangi and other 45 previously published spider mitogenome sequences, including that of Argyroneta aquatica. We found a unique transposition of trnL2 and trnN genes in Desis jiaxiangi. Our robust phylogenetic topology clearly deciphered the evolutionary relationships between Desis jiaxiangi and Argyroneta aquatica as well as other spiders. We dated the divergence of Desis jiaxiangi and Argyroneta aquatica to the late Cretaceous at ~ 98 Ma. Our selection analyses detected a positive selection signal in the nd4 gene of the aquatic branch comprising both Desis jiaxiangi and Argyroneta aquatica. Surprisingly, Pirata subpiraticus, Hypochilus thorelli, and Argyroneta aquatica each had a higher Ka/Ks value in the 13 PCGs dataset among 46 taxa with complete mitogenomes, and these three species also showed positive selection signal in the nd6 gene. Conclusions Our finding of the unique transposition of trnL2 and trnN genes indicates that these genes may have experienced rearrangements in the history of intertidal spider evolution. The positive selection signals in the nd4 and nd6 genes might enable a better understanding of the spider metabolic adaptations in relation to different environments. Our construction of a novel mitogenome for the intertidal spider thus sheds light on the evolutionary history of spiders and their mitogenomes.

Abstract Although there are various Conus species with publicly available transcriptome and proteome data, no genome assembly has been reported yet. Here, using Chinese tubular cone snail (C. betulinus) as a representative, we sequenced and assembled the first Conus genome with original identification of 133 genome-widely distributed conopeptide genes. After integration of our genomics, transcriptomics, and peptidomics data in the same species, we established a primary genetic central dogma of diverse conopeptides, assuming a rough number ratio of ~1:1:1:10s for the total genes: transcripts: proteins: post-translationally modified peptides. This ratio may be special for this worm-hunting Conus species, due to the high diversity of various Conus genomes and the big number ranges of conopeptide genes, transcripts, and peptides in previous reports of diverse Conus species. Only a fraction (45.9%) of the identified conotopeptide genes from our achieved genome assembly are transcribed with transcriptomic evidence, and few genes individually correspond to multiple transcripts possibly due to intraspecies or mutation-based variances. Variable peptide processing at the proteomic level, generating a big diversity of venom conopeptides with alternative cleavage sites, post-translational modifications, and N-/C-terminal truncations, may explain how the 133 genes and ~123 transcripts can generate thousands of conopeptides in the venom of individual C. betulinus. We also predicted many conopeptides with high stereostructural similarities to the putative analgesic ω-MVIIA, addiction therapy AuIB and insecticide ImI, suggesting that our current genome assembly for C. betulinus is a valuable genetic resource for high-throughput prediction and development of potential pharmaceuticals.

As an important hormone, melatonin participates in endocrine regulation of diverse functions in vertebrates. Its biosynthesis is catalyzed by four cascaded enzymes, among them, arylalkylamine N-acetyltransferase (AANAT) is the most critical one. Although only single aanat gene has been identified in most groups of vertebrates, researchers including us have determined that fish have the most diverse of aanat genes (aanat1a, aanat1b, and aanat2), playing various potential roles such as seasonal migration, amphibious aerial vision, and cave or deep-sea adaptation. With the rapid development of genome and transcriptome sequencing, more and more putative sequences of fish aanat genes are going to be available. Related phylogeny and functional investigations will enrich our understanding of AANAT functions in various fish species.

BackgroundThe novel coronavirus disease 2019 (COVID-19) pandemic causes great disruption to cancer care services, which might bring about psychological problems and further lower both physical and mental life quality in cancer patients. Until now, very few studies focused on the psychological distress of patients with advanced melanoma before or during the epidemic. This study aimed to elucidate the fear of progression (FoP), anxiety, depression, and related independent predictors in patients with advanced melanoma during the COVID-19 outbreak.MethodsTwo hundred and seventy-three patients with unresectable stage III or metastatic melanoma were recruited from February 2020 to November 2021, and completed the Fear of Progression Questionnaire-Short Form (FoP-Q-SF), State Trait Anxiety Inventory (STAI-6), and Patient Health Questionnaire (PHQ-9).ResultsOne hundred and seventy-four (64.7%) patients experienced heighted FoP (FoP-Q-SF: 39.9 ± 11.0), 198 (72.5%) patients reported elevated anxiety (STAI-6: 13.1 ± 3.0), and 62 (22.7%) patients had increased depression (PHQ-9: 6.4 ± 6.1). In multivariate analysis, illness duration (OR = 0.987 for FoP; OR = 0.984 for depression), cancer stage (OR = 14.394 for anxiety) and disease progression (OR = 1.960 for FoP; OR = 23.235 for anxiety; OR = 1.930 for depression) were independent predictors for FoP, anxiety or depression. Additionally, the high levels of FoP, anxiety and depression were significantly positive correlated with each other (r = 0.466 for FoP and anxiety; r = 0.382 for FoP and depression; r = 0.309 for anxiety and depression).ConclusionOur study indicates that FoP, anxiety and depression are persisting among patients with advanced melanoma in the COVID-19 and post-COVID-19 era. Effective psycho-oncological interventions are needed for melanoma patients with psychological distress during the ongoing COVID-19 pandemic.

Resveratrol is a natural active ingredient found in many plants and has a protective effect on cardiovascular diseases. However, it has not been reported that whether resveratrol can inhibit calcific aortic valve disease(CAVD). In this study, we want to explore the effect and mechanism of resveratrol on the osteogenic differentiation of valve interstitial cells（VICs）.We first found that the expression of osteogenic markers in calcified human aortic valve. Western blotting, ALP staining and Alizarin red S staining showed that under osteogenic medium (OM) treatment, resveratrol can inhibit VICs from expressing osteogenic markers, reduce ALP content and calcium Nodule formation. Mechanistically, resveratrol can inhibit the activation of the AKT/Smad1/5/8 pathway induced by OM. We selected the AKT pathway for verification and determined that resveratrol inhibited the osteogenic differentiation of VICs by inhibiting the AKT pathway. Collectively, resveratrol may become a potential natural drug for the treatment of CAVD.

Plants are a rich source for bioactive compounds. However, plant extracts can harbor a mixture of bioactive molecules that promote divergent phenotypes and potentially have confounding effects in bioassays. Even with further purification and identification, target deconvolution can be challenging. Corynoline and acetylcorynoline, are phytochemicals that were previously isolated through a screen for compounds able to induce mitotic arrest and polyploidy in oncogene expressing retinal pigment epithelial (RPE) cells. Here, we shed light on the mechanism by which these phytochemicals can attack human cancer cells. Mitotic arrest was coincident to the induction of centrosome amplification and declustering, causing multi-polar spindle formation. Corynoline was demonstrated to have true centrosome declustering activity in a model where A549 cells were chemically induced to have more than a regular complement of centrosomes. Corynoline could inhibit the centrosome clustering required for pseudo-bipolar spindle formation in these cells. The activity of AURKB, but not AURKA or polo-like kinase 4, was diminished by corynoline. It only partially inhibited AURKB, so it may be a partial antagonist or corynoline may work upstream on an unknown regulator of AURKB activity or localization. Nonetheless, corynoline and acetylcorynoline inhibited the viability of a variety of human cancer derived cell lines. These phytochemicals could serve as prototypes for a next-generation analog with improved potency, selectivity or in vivo bioavailability. Such an analog could be useful as a non-toxic component of combination therapies where inhibiting the chromosomal passenger protein complex is desired.