Integrative omics analysis elucidates the genetic basis underlying seed weight and oil content in soybean.

Synergistic optimization of key agronomic traits by traditional breeding has dramatically enhanced crop productivity in the past decades. However, the genetic basis underlying coordinated regulation of yield- and quality-related traits remains poorly understood. Here, we dissected the genetic architectures of seed weight and oil content by combining genome-wide association studies (GWAS) and transcriptome-wide association studies (TWAS) using 421 soybean (Glycine max) accessions. We identified 26 and 33 genetic loci significantly associated with seed weight and oil content by GWAS, respectively, and detected 5,276 expression quantitative trait loci (eQTLs) regulating expression of 3,347 genes based on population transcriptomes. Interestingly, a gene module (IC79), regulated by two eQTL hotspots, exhibited significant correlation with both seed weigh and oil content. Twenty-two candidate causal genes for seed traits were further prioritized by TWAS, including Regulator of Weight and Oil of Seed 1 (GmRWOS1), which encodes a sodium pump protein. GmRWOS1 was verified to pleiotropically regulate seed weight and oil content by gene knockout and overexpression. Notably, allelic variations of GmRWOS1 were strongly selected during domestication of soybean. This study uncovers the genetic basis and network underlying regulation of seed weight and oil content in soybean and provides a valuable resource for improving soybean yield and quality by molecular breeding.

A neural pathway for social modulation of spontaneous locomotor activity (SoMo-SLA) in Drosophila.

Social enrichment or social isolation affects a range of innate behaviors, such as sex, aggression, and sleep, but whether there is a shared mechanism is not clear. Here, we report a neural mechanism underlying social modulation of spontaneous locomotor activity (SoMo-SLA), an internal-driven behavior indicative of internal states. We find that social enrichment specifically reduces spontaneous locomotor activity in male flies. We identify neuropeptides Diuretic hormone 44 (DH44) and Tachykinin (TK) to be up- and down-regulated by social enrichment and necessary for SoMo-SLA. We further demonstrate a sexually dimorphic neural circuit, in which the male-specific P1 neurons encoding internal states form positive feedback with interneurons coexpressing doublesex (dsx) and Tk to promote locomotion, while P1 neurons also form negative feedback with interneurons coexpressing dsx and DH44 to inhibit locomotion. These two opposing neuromodulatory recurrent circuits represent a potentially common mechanism that underlies the social regulation of multiple innate behaviors.

Asexuality in Drosophila juvenile males is organizational and independent of juvenile hormone.

Sexuality is generally prevented in newborns and arises with organizational rewiring of neural circuitry and optimization of fitness for reproduction competition. Recent studies reported that sex circuitry in Drosophila melanogaster is developed in juvenile males but functionally inhibited by juvenile hormone (JH). Here, we find that the fly sex circuitry, mainly expressing the male-specific fruitless (fruM ) and/or doublesex (dsx), is organizationally undeveloped and functionally inoperative in juvenile males. Artificially activating all fruM neurons induces substantial courtship in solitary adult males but not in juvenile males. Synaptic transmissions between major courtship regulators and all dsx neurons are strong in adult males but either weak or undetectable in juvenile males. We further find that JH does not inhibit male courtship in juvenile males but instead promotes courtship robustness in adult males. Our results indicate that the transition to sexuality from juvenile to adult flies requires organizational rewiring of neural circuitry.

The doublesex gene regulates dimorphic sexual and aggressive behaviors in Drosophila.

Most animal species display dimorphic sexual behaviors and male-biased aggressiveness. Current models have focused on the male-specific product from the fruitless (fruM) gene, which controls male courtship and male-specific aggression patterns in fruit flies, and describe a male-specific mechanism underlying sexually dimorphic behaviors. Here we show that the doublesex (dsx) gene, which expresses male-specific DsxM and female-specific DsxF transcription factors, functions in the nervous system to control both male and female sexual and aggressive behaviors. We find that Dsx is not only required in central brain neurons for male and female sexual behaviors, but also functions in approximately eight pairs of male-specific neurons to promote male aggressiveness and approximately two pairs of female-specific neurons to inhibit female aggressiveness. DsxF knockdown females fight more frequently, even with males. Our findings reveal crucial roles of dsx, which is broadly conserved from worms to humans, in a small number of neurons in both sexes to establish dimorphic sexual and aggressive behaviors.

The emergence of machine learning force fields in drug design.

In the field of molecular simulation for drug design, traditional molecular mechanic force fields and quantum chemical theories have been instrumental but limited in terms of scalability and computational efficiency. To overcome these limitations, machine learning force fields (MLFFs) have emerged as a powerful tool capable of balancing accuracy with efficiency. MLFFs rely on the relationship between molecular structures and potential energy, bypassing the need for a preconceived notion of interaction representations. Their accuracy depends on the machine learning models used, and the quality and volume of training data sets. With recent advances in equivariant neural networks and high-quality datasets, MLFFs have significantly improved their performance. This review explores MLFFs, emphasizing their potential in drug design. It elucidates MLFF principles, provides development and validation guidelines, and highlights successful MLFF implementations. It also addresses potential challenges in developing and applying MLFFs. The review concludes by illuminating the path ahead for MLFFs, outlining the challenges to be overcome and the opportunities to be harnessed. This inspires researchers to embrace MLFFs in their investigations as a new tool to perform molecular simulations in drug design.

DNA methylation remodeled amino acids biosynthesis regulates flower senescence in carnation (Dianthus caryophyllus).

Dynamic DNA methylation regulatory networks are involved in many biological processes. However, how DNA methylation patterns change during flower senescence and their relevance with gene expression and related molecular mechanism remain largely unknown. Here, we used whole genome bisulfite sequencing to reveal a significant increase of DNA methylation in the promoter region of genes during natural and ethylene-induced flower senescence in carnation (Dianthus caryophyllus L.), which was correlated with decreased expression of DNA demethylase gene DcROS1. Silencing of DcROS1 accelerated while overexpression of DcROS1 delayed carnation flower senescence. Moreover, among the hypermethylated differentially expressed genes during flower senescence, we identified two amino acid biosynthesis genes, DcCARA and DcDHAD, with increased DNA methylation and reduced expression in DcROS1 silenced petals, and decreased DNA methylation and increased expression in DcROS1 overexpression petals, accompanied by decreased or increased amino acids content. Silencing of DcCARA and DcDHAD accelerates carnation flower senescence. We further showed that adding corresponding amino acids could largely rescue the senescence phenotype of DcROS1, DcCARA and DcDHAD silenced plants. Our study not only demonstrates an essential role of DcROS1-mediated remodeling of DNA methylation in flower senescence but also unravels a novel epigenetic regulatory mechanism underlying DNA methylation and amino acid biosynthesis during flower senescence.

Histone H3K4 methyltransferase DcATX1 promotes ethylene induced petal senescence in carnation.

Petal senescence is controlled by a complex regulatory network. Epigenetic regulation like histone modification influences chromatin state and gene expression. However, the involvement of histone methylation in regulating petal senescence remains poorly understood. Here, we found that the trimethylation of histone H3 at Lysine 4 (H3K4me3) is increased during ethylene-induced petal senescence in carnation (Dianthus caryophyllus L.). H3K4me3 levels were positively associated with the expression of transcription factor DcWRKY75, ethylene biosynthetic genes 1-aminocyclopropane-1-carboxylic acid (ACC) synthase (DcACS1), and ACC oxidase (DcACO1), and senescence associated genes (SAGs) DcSAG12 and DcSAG29. Further, we identified that carnation ARABIDOPSIS HOMOLOG OF TRITHORAX1 (DcATX1) encodes a histone lysine methyltransferase which can methylate H3K4. Knockdown of DcATX1 delayed ethylene-induced petal senescence in carnation, which was associated with the down-regulated expression of DcWRKY75, DcACO1, and DcSAG12, whereas overexpression of DcATX1 exhibited the opposite effects. DcATX1 promoted the transcription of DcWRKY75, DcACO1, and DcSAG12 by elevating the H3K4me3 levels within their promoters. Overall, our results demonstrate that DcATX1 is a H3K4 methyltransferase that promotes the expression of DcWRKY75, DcACO1, DcSAG12 and potentially other downstream target genes by regulating H3K4me3 levels, thereby accelerating ethylene-induced petal senescence in carnation. This study further indicates that epigenetic regulation is important for plant senescence processes.

Altered chromatin architecture and gene expression during polyploidization and domestication of soybean.

Polyploidy or whole-genome duplication (WGD) is widespread in plants and is a key driver of evolution and speciation, accompanied by rapid and dynamic changes in genomic structure and gene expression. The 3D structure of the genome is intricately linked to gene expression, but its role in transcription regulation following polyploidy and domestication remains unclear. Here, we generated high-resolution (∼2 kb) Hi-C maps for cultivated soybean (Glycine max), wild soybean (Glycine soja), and common bean (Phaseolus vulgaris). We found polyploidization in soybean may induce architecture changes of topologically associating domains and subsequent diploidization led to chromatin topology alteration around chromosome-rearrangement sites. Compared with single-copy and small-scale duplicated genes, WGD genes displayed more long-range chromosomal interactions and were coupled with higher levels of gene expression and chromatin accessibilities but void of DNA methylation. Interestingly, chromatin loop reorganization was involved in expression divergence of the genes during soybean domestication. Genes with chromatin loops were under stronger artificial selection than genes without loops. These findings provide insights into the roles of dynamic chromatin structures on gene expression during polyploidization, diploidization, and domestication of soybean.

A regional genomic surveillance program is implemented to monitor the occurrence and emergence of SARS-CoV-2 variants in Yubei District, China.

BACKGROUND: In December 2022, Chongqing experienced a significant surge in coronavirus disease 2019 (COVID-19) epidemic after adjusting control measures in China. Given the widespread immunization of the population with the BA.5 variant, it is crucial to actively monitor severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variant evolution in Chongqing's Yubei district. METHODS: In this retrospective study based on whole genome sequencing, we collected oropharyngeal and nasal swab of native COVID-19 cases from Yubei district between January to May 2023, along with imported cases from January 2022 to January 2023. Through second-generation sequencing, we generated a total of 578 genomes. RESULTS: Phylogenetic analyses revealed these genomes belong to 47 SARS-CoV-2 Pango lineages. BA.5.2.48 was dominant from January to April 2023, rapidly replaced by XBB* variants from April to May 2023. Bayesian Skyline Plot reconstructions indicated a higher evolutionary rate (6.973 × 10-4 subs/site/year) for the XBB.1.5* lineage compared to others. The mean time to the most recent common ancestor (tMRCA) of BA.5.2.48* closely matched BA.2.75* (May 27, 2022). Using multinomial logistic regression, we estimated growth advantages, with XBB.1.9.1 showing the highest growth advantage (1.2, 95% HPI:1.1-1.2), followed by lineage FR.1 (1.1, 95% HPI:1.1-1.2). CONCLUSIONS: Our monitoring reveals the rapid replacement of the previously prevalent BA.5.2.48 variant by XBB and its sub-variants, underscoring the ineffectiveness of herd immunity and breakthrough BA.5 infections against XBB variants. Given the ongoing evolutionary pressure, sustaining a SARS-CoV-2 genomic surveillance program is imperative.

Systemic lupus erythematosus with chronic persistent intracranial hypertension: A case report.

OBJECTIVE: The aim is to investigate the clinical characteristics of systemic lupus erythematosus with intracranial hypertension. METHODS: The clinical characteristics of one case of systemic lupus erythematosus with chronic persistent intracranial hypertension were analyzed, and related literature was reviewed by searching Medline and Wanfang databases. RESULTS: Intracranial hypertension in SLE patients may occur at the onset or during the course of the disease. Our patient was diagnosed with IH 3 years after the onset of SLE. Headache and papilledema were the most common symptoms of intracranial hypertension, followed by nausea or vomiting, vision changes, and cerebral palsy. Our patient had a headache and cranial hypertension that lasted for years, but no papilledema was found. Corticosteroid is currently the mainstay of the treatment of IIH in patients with SLE, and immunosuppressive agents, acetazolamide, intravenous mannitol and furosemide are also used. However, our patient did not respond to these treatments and presents the characteristics of chronic persistent intracranial hypertension. CONCLUSION: Systemic lupus erythematosus with intracranial hypertension is a rare manifestation of SLE, which is not completely parallel to SLE activity. Headache and papilledema were the most common presenting symptoms. Different from previous reported cases, our patient had poor response to treatments, showing chronic and persistent characteristics.

Characteristics of CD4-1 gene and its immune responses against Aeromonas veronii infection by activating NF-κB signaling in Qihe crucian carp Carassius auratus.

CD4-1 found in bony fish contains four extracellular immunoglobulin (Ig)-like domains similar to that of mammalian CD4, which is crucial for the activation of CD4+ helper T-cell. However, there is limited knowledge regarding the molecular markers, immune functions and regulation mechanism of CD4-1 in teleosts due to their vast diversity. In this study, we cloned and characterized two isoforms of Qihe crucian carp CD4-1, designated as CaCD4-1.1 and CaCD4-1.2. We further explored their expression responses upon stimulation with Aeromonas veronii, and the regulation of their immune responses against A. veronii by NF-κB. The ORF of CaCD4-1.1 and CaCD4-1.2 cDNA encoded 477 and 466 amino acids, respectively. Both proteins contained seven conserved cysteine residues in the extracellular domain, and a CCC motif in their cytoplasm, respectively. However, CaCD4-1.1 exhibited a relatively limited similarity with CaCD4-1.2 in the ectodomain. The quantitative real-time polymerase chain reaction (qRT-PCR) analysis revealed that the mRNA expression of CaCD4-1.1 and CaCD4-1.2 exhibited differential constitutive expression across all examined tissues. Furthermore, the expression level of CD4-1.2 was higher than that of CD4-1.1 in the gills, head kidney, and spleen of Qihe crucian carp subjected to A. veronii challenge, while it was lower in the trunk kidney. Inhibition of NF-κB activity resulted in a decrease in the expression levels of CD4-1.1 and CD4-1.2 mRNA in the gill, while inducing an increase in expression levels in the spleen, in accordance with the observed ultrastructural changes in both organs. Interestingly, the impact of NF-κB on the mRNA expression level of CD4-1.1 appears to be stronger than that of CD4-1.2. Our results suggest that CaCD4-1.1 and CaCD4-1.2 could be expressed on T cells and antigen-sampling cells that exhibit similar characteristics to mammalian M cells, respectively, and differentially regulated by NF-κB in adaptive immune responses against bacterial infection. This research contributes to a better understanding of the crucial role of CD4-1 in the immune response of Qihe crucian carp and provide novel insights for the prevention and treatment of fish diseases in aquaculture.

MiR-214_L-1R+4 regulate gossypol-induced immune response through MyD88-dependent signaling pathway in Cyprinus carpio.

MicroRNAs (miRNAs) have been demonstrated to act as crucial modulators with considerable impacts on the immune system. Cottonseed meal is often used as a protein source in aqua feed, cottonseed meal contains gossypol, which is harmful to animals. However, there is a lack of research on the role of miRNAs in fish exposed to gossypol stress. To determine the regulatory effects of miRNAs on gossypol toxicity, Cyprinus carpio were given to oral administration of 20 mg/kg gossypol for 7 days, and the gossypol concentration in the tissues was tested. Then, we detected spleen index, histology, immune enzyme activities of fish induced by gossypol. The results of miRNA sequencing revealed 8 differentially expressed miRNAs in gossypol group, and miR-214_L-1R+4 was found involved in immune response induced by gossypol. The potential targets of miR-214_L-1R+4 were predicted, and found a putative miR-214_L-1R+4 binding site in the 3'UTR of MyD88a. Furthermore, dual-luciferase reporter assays displayed miR-214_L-1R+4 decreased MyD88a expression through binding to the 3'UTR of MyD88a. Moreover, miR-214_L-1R+4 antagomir were intraperitoneally administered to C. carpio, down-regulated miR-214_L-1R+4 could increase MyD88a expression, as well as inflammatory cytokines and anti-inflammatory cytokines expression. These findings revealed that miR-214_L-1R+4 via the MyD88-dependent signaling pathway modulate the immune response to gossypol in C. carpio spleen.

Assessing the economic ripple effect of flood disasters in light of the recovery process: Insights from an agent-based model.

To assess the economic ripple effect, this study integrates agent-based modeling (ABM) with a multiregional input-output (MRIO) table to develop an assessment model that considers capacity recovery process. The intermediate and final demands in the MRIO table are used to describe the agents' interdependence. Survival analysis is used to construct capacity rate curves. By defining the first- and second-order ripple effects, ABM is used to capture the ripple process in days. To conduct a case study, the service and retail sectors in Enshi in Hubei, China, are selected as disaster-affected sectors (they were severely affected by the July 17, 2020 flood disaster). The main findings are as follows: (1) With the first-order ripple effect, the losses caused by service and retail are concentrated within Enshi. Enshi's final demand, construction, and raw materials manufacturing sectors as well as Wuhan's construction sector are seriously affected. (2) With the second-order ripple effect, the losses caused by the service and retail sectors expand, forming a prominent industrial ripple chain: "service (retail)-raw materials manufacturing-construction." (3) The direct and indirect losses caused by the service sector are more significant than those caused by the retail sector. However, the loss ratio of the service sector is smaller than that of the retail sector because of its sound industrial structure and strong resilience. Hence, the indirect losses caused by different sectors are not entirely determined by their direct losses; instead, they are also related to the degree of perfection of the structures of different sectors.

Template-Free Synthesis of Phosphorus-Doped g-C3 N4 Micro-Tubes with Hierarchical Core-Shell Structure for High-Efficient Visible Light Responsive Catalysis.

This work reports a new form of tubular g-C3 N4 that is featured with a hierarchical core-shell structure introduced with phosphorous elements and nitrogen vacancies. The core is self-arranged with randomly stacked g-C3 N4 ultra-thin nanosheets along the axial direction. This unique structure significantly benefits electron/hole separation and visible-light harvesting. A superior performance for the photodegradation of rhodamine B and tetracycline hydrochloride is demonstrated under low intensity visible light. This photocatalyst also exhibits an excellent hydrogen evolution rate (3631 µmol h-1 g-1 ) under visible light. Realizing this structure just requires the introduction of phytic acid into the solution of melamine and urea during hydrothermal treatment. In this complex system, phytic acid plays as the electron donor to stabilize melamine/cyanuric acid precursor via coordination interaction. Calcination at 550 °C directly renders the transformation of precursor into such hierarchical structure. This process is facile and shows the strong potential toward mass production for real applications.

An insertion of transposon in DcNAP inverted its function in the ethylene pathway to delay petal senescence in carnation (Dianthus caryophyllus L.).

Petal senescence is the final stage of flower development. Transcriptional regulation plays key roles in this process. However, whether and how post-transcriptional regulation involved is still largely unknown. Here, we identified an ethylene-induced NAC family transcription factor DcNAP in carnation (Dianthus caryophyllus L.). One allele, DcNAP-dTdic1, has an insertion of a dTdic1 transposon in its second exon. The dTdic1 transposon disrupts the structure of DcNAP and causes alternative splicing, which transcribes multiple domain-deleted variants (DcNAP2 and others). Conversely, the wild type allele DcNAP transcribes DcNAP1 encoding an intact NAC domain. Silencing DcNAP1 delays and overexpressing DcNAP1 accelerates petal senescence in carnation, while silencing and overexpressing DcNAP2 have the opposite effects, respectively. Further, DcNAP2 could interact with DcNAP1 and interfere the binding and activation activity of DcNAP1 to the promoters of its downstream target ethylene biosynthesis genes DcACS1 and DcACO1. Lastly, ethylene signalling core transcriptional factor DcEIL3-1 can activate the expression of DcNAP1 and DcNAP2 in the same way by binding their promoters. In summary, we discovered a novel mechanism by which DcNAP regulates carnation petal senescence at the post-transcriptional level. It may also provide a useful strategy to manipulate the NAC domains of NAC transcription factors for crop genetic improvement.

DEL-1, as an anti-neutrophil transepithelial migration molecule, inhibits airway neutrophilic inflammation in asthma.

BACKGROUND: Neutrophil migration into the airways is a key process in neutrophilic asthma. Developmental endothelial locus-1 (DEL-1), an extracellular matrix protein, is a neutrophil adhesion inhibitor that attenuates neutrophilic inflammation. METHODS: Levels of DEL-1 were measured in exhaled breath condensate (EBC) and serum in asthma patients by ELISA. DEL-1 modulation of neutrophil adhesion and transepithelial migration was examined in a co-culture model in vitro. The effects of DEL-1-adenoviral vector-mediated overexpression on ovalbumin/lipopolysaccharide (OVA/LPS)-induced neutrophilic asthma were studied in mice in vivo. RESULTS: DEL-1 was primarily expressed in human bronchial epithelial cells and was decreased in asthma patients. Serum DEL-1 concentrations were reduced in patients with severe asthma compared with normal subjects (567.1 ± 75.3 vs. 276.8 ± 29.36 pg/mL, p < .001) and were negatively correlated to blood neutrophils (r = -0.2881, p = .0384) and neutrophil-to-lymphocyte ratio (NLR) (r = -0.5469, p < .0001). DEL-1 concentrations in the EBC of severe asthmatic patients (113.2 ± 8.09 pg/mL) were also lower than normal subjects (193.0 ± 7.61 pg/mL, p < .001) and were positively correlated with the asthma control test (ACT) score (r = 0.3678, p = .0035) and negatively related to EBC IL-17 (r = -0.3756, p = .0131), myeloperoxidase (MPO) (r = -0.5967, p = .0055), and neutrophil elastase (NE) (r = -0.5488, p = .0009) expression in asthma patients. Neutrophil adhesion and transepithelial migration in asthma patients were associated with LFA-1 binding to ICAM-1 and inhibited by DEL-1. DEL-1 mRNA and protein expression in human bronchial epithelial cells were regulated by IL-17. Exogenous DEL-1 inhibited IL-17-enhanced neutrophil adhesion and migration. DEL-1 expression was decreased while neutrophil infiltration was increased in the airway of a murine model of neutrophilic asthma. This was prevented by DEL-1 overexpression. CONCLUSIONS: DEL-1 down-regulation leads to increased neutrophil migration across bronchial epithelial cells and is associated with neutrophilic airway inflammation in asthma.

Hydrogen Peroxide-Activated Nitric Oxide-Releasing Vancomycin-Loaded Electrostatic Complexation for Efficient Elimination of Methicillin-Resistant Staphylococcus aureus Abscesses.

The treatment of subcutaneous abscesses has been greatly hindered due to the spread of drug-resistant strains such as methicillin-resistant Staphylococcus aureus (MRSA). Thus, alternative strategies are highly desired to complement conventional antibiotic therapies and surgical intervention. As one of such strategies, applications of nitric oxide (NO) have shown great potential in the treatment of bacteria-induced subcutaneous abscesses by improving the efficacy of many therapeutic methods. However, it is extremely challenging to achieve precise delivery and controlled release because of its gaseous nature. In the present study, an effective strategy was reported in which on demand hydrogen peroxide (H2O2)-activated nitric oxide-releasing vancomycin (Van)-loaded electrostatic complexation (Lipo/Van@Arg) was fabricated. In this system, Van was encapsulated into a negative-charged DSPG/Chol liposome (Lipo/Van) and electrostatically bound with the positive-charged l-arginine (l-Arg). As expected, Lipo/Van@Arg exhibited superior bacterial binding and biofilm penetration abilities. After being in the interior of the biofilms, Lipo/Van@Arg could be triggered by the endogenous H2O2 and effectively release NO. The released NO could exhibit combined antibacterial and biofilm eradication effects with Van. Moreover, an in vivo evaluation using a BALB/c mouse model of subcutaneous abscesses indicated that the combination treatment of NO and Van based on Lipo/Van@Arg could effectively eliminate MRSA from the abscesses, thereby preventing abscess recurrence. In summary, the Lipo/Van@Arg system developed in this study realized controlled delivery and precise release of NO, which had significant clinical implications in the efficient treatment of abscesses.

Characterization of CD3γ/δ gene and its immune response in Qihe crucian carp Carassius auratus after challenged by Aeromonas veronii and Poly(I:C).

CD3γ/δ found in non-mammalian vertebrates is a CD3 homolog with structural characteristics similar to both mammalian CD3γ and CD3δ, and plays important roles in T cell recognization and immune response in fish. In this study, the full-length of CD3γ/δ from Qihe crucian carp (named CaCD3γ/δ) was cloned and characterized, then the expression response profiles and potential immune functions was explored after Aeromonas veronii and Poly(I:C) challenge. The results showed that the full-length of CaCD3γ/δ was 819 bp including a 5'-UTR of 141 bp, a 3'-UTR of 168 bp, and an ORF of 510 bp encoding a putative 169-aa protein with an estimated MW of 18.71 kD and a theoretical pI of 8.77. The protein sequence of CaCD3γ/δ contained a Leu-Leu and a CXXXC motif in the extracellular domain, and an ITAM and a Leu-Ile motif in the cytoplasm, and a residue of Asn in the transmembrane. CaCD3γ/δ was constitutively expressed in the spleen, liver, gill, and blood of Qihe crucian carp. After the carp were challenged with Poly(I:C) and Aeromonas veronii, the mRNA expression levels of CaCD3γ/δ were significantly changed in the spleen, head kidney, intestine and gill, according to the results of qPCR. However, compared with A. veronii, Poly(I:C) challenge can rapidly induce the CaCD3γ/δ expression levels in head kidney, intestine and spleen, which suggested CaCD3γ/δ may be differentially modulated by different pathogens. Moreover, the results of immunohistochemical analysis showed that the CaCD3γ/δ+ secreted cells in the spleen and gills of Qihe crucian were increased after challenged with Poly(I:C), as well as the spleen challenged with A. veronii, but at different levels. Combined with the fact that vascular congestion, necrosis of parenchymal cells, and inflammatory cells including lymphocytes infiltration were also observed in the gill and spleen of Qihe crucian carp treated with A. veronii and Poly(I:C) revealed by pathological analysis, it was predicted that CaCD3γ/δ+ T lymphocytes may participated in the immune response against pathogens. This study will contribute to understand the important role of CaCD3γ/δ+ T lymphocytes in the immune response of Qihe crucian carp, and provide new insights for the prevention and treatment of the diseases of Qihe crucian carp.

Two CcCCL19bs orchestrate an antibacterial immune response in Yellow River carp (Cyprinus carpio haematopterus).

Chemokines are a group of chemotactic cytokines with an essential role in homeostasis as well as immunity via specific G protein-coupled receptors and atypical receptors. In our study, two Yellow River carp (Cyprinus carpio haematopterus) CCL19b genes (CcCCL19bs), tentatively named CcCCL19b_a and CcCCL19b_b, were cloned. The open reading frames (ORFs) of CcCCL19b_a and CcCCL19b_b were both 333 bp that encoded a 12 kDa protein with 110 amino acid residues. CcCCL19bs contained a signal peptide and a SCY domain with four typical conserved cysteine residues. The two CcCCL19b proteins shared high similarities with each other in both secondary and three-dimensional structure. Phylogenetic analysis showed that CcCCL19bs and other CCL19bs from tetraploid cyprinid fish were clustered into one clade. CcCCL19bs were highly expressed in gill and intestine in healthy fish, and a significant up-regulation of gene expression after Aeromonas hydrophila infection and poly(I:C) stimulation was observed in gill, liver, and head kidney. Furthermore, chemotaxis and antibacterial activity of CcCCL19bs were studied. The results indicated that recombinant CcCCL19b_a and CcCCL19b_b protein (rCcCCL19b_a and rCcCCL19b_b) exhibited significant attraction to primary head kidney leukocytes (HKLs). Meanwhile, both of rCcCCL19bs could promote the proliferation of HKLs, and significantly up-regulate the expressions of IL-1ß, CCR7, and IL-6, and down-regulate the expression of IL-10 in primary HKLs. In vitro, rCcCCL19bs could bind and aggregate A. hydrophila and Staphylococcus aureus. The rCcCCL19bs exhibited significant antibacterial activity against A. hydrophila, but not S. aureus. Moreover, they inhibited the growth of A. hydrophila and S. aureus. In vivo, overexpression of CcCCL19bs contributed to the bacterial clearance. These studies suggested that CcCCL19bs orchestrate an antibacterial immune response.

Functional roles of CcGSDMEa-like in common carp (Cyprinus carpio) after Aeromonas hydrophila infection.

GSDMs could punch holes in cell membrane and participate in the immune response to bacterial infections. In current study, the molecular and structural characteristics of CcGSDMEa-like were analyzed, and the role of CcGSDMEa-like in the inflammatory response against Aeromonas hydrophila was studied. The results showed that the CcGSDMEa-like shared the conserved structural characteristics with GSDMEs of other teleosts. The CcGSDMEa-like mRNA and protein expression levels were significantly affected by A. hydrophila challenge. When the CcGSDMEa-like was overexpressed, the expression of CcIL-1ß were significantly increased in fish and EPC cells, and bacterial contents were significantly decreased in fish tissues. While, when the CcGSDMEa-like was knocked down, the expression and secretion of CcIL-1ß were significantly decreased in vivo and in vitro, and the bacterial contents were increased in vivo after A. hydrophila infection 12 h and 24 h. In brief, CcGSDMEa-like could regulate the content of bacteria in fish through mediating the expression and secretion of CcIL-1ß. Bactericidal assay and cytotoxicity assay showed that CcGSDMEa-like had no bactericidal activity to Escherichia coli, and did not disrupt cytomembrane integrity of HEK293T cells. This study suggested that CcGSDMEa-like could play roles in the antibacterial and inflammatory processes in fish.