DDX5 inhibits inflammation by modulating m6A levels of TLR2/4 transcripts during bacterial infection.

DExD/H-box helicases are crucial regulators of RNA metabolism and antiviral innate immune responses; however, their role in bacteria-induced inflammation remains unclear. Here, we report that DDX5 interacts with METTL3 and METTL14 to form an m6A writing complex, which adds N6-methyladenosine to transcripts of toll-like receptor (TLR) 2 and TLR4, promoting their decay via YTHDF2-mediated RNA degradation, resulting in reduced expression of TLR2/4. Upon bacterial infection, DDX5 is recruited to Hrd1 at the endoplasmic reticulum in an MyD88-dependent manner and is degraded by the ubiquitin-proteasome pathway. This process disrupts the DDX5 m6A writing complex and halts m6A modification as well as degradation of TLR2/4 mRNAs, thereby promoting the expression of TLR2 and TLR4 and downstream NF-κB activation. The role of DDX5 in regulating inflammation is also validated in vivo, as DDX5- and METTL3-KO mice exhibit enhanced expression of inflammatory cytokines. Our findings show that DDX5 acts as a molecular switch to regulate inflammation during bacterial infection and shed light on mechanisms of quiescent inflammation during homeostasis.

Identification and Characterization of chCR2, a Protein That Binds Chicken Complement Component 3d.

Complement receptor type 2 (CR2) is an important membrane molecule expressed on B cells and follicular dendritic cells. Human CR2 has been shown to play a critical role in bridging the innate complement-mediated immune response with adaptive immunity by binding complement component 3d (C3d). However, the chicken CR2 (chCR2) gene has not been identified or characterized. In this study, unannotated genes that contain short consensus repeat (SCR) domains were analyzed based on RNA sequencing data for chicken bursa lymphocytes, and a gene with >80% homology to CR2 from other bird species was obtained. The gene consisted of 370 aa and was much smaller than the human CR2 gene because 10-11 SCRs were missing. The gene was then demonstrated as a chCR2 that exhibited high binding activity to chicken C3d. Further studies revealed that chCR2 interacts with chicken C3d through a binding site in its SCR1-4 region. An anti-chCR2 mAb that recognizes the epitope 258CKEISCVFPEVQ269 was prepared. Based on the anti-chCR2 mAb, the flow cytometry and confocal laser scanning microscopy experiments confirmed that chCR2 was expressed on the surface of bursal B lymphocytes and DT40 cells. Immunohistochemistry and quantitative PCR analyses further indicated that chCR2 is predominantly expressed in the spleen, bursa, and thymus, as well as in PBLs. Additionally, the expression of chCR2 varied according to the infectious bursal disease virus infection status. Collectively, this study identified and characterized chCR2 as a distinct immunological marker in chicken B cells.

Interlayer Fermi Polarons of Excited Exciton States in Quantizing Magnetic Fields.

The study of exciton polarons has offered profound insights into the many-body interactions between bosonic excitations and their immersed Fermi sea within layered heterostructures. However, little is known about the properties of exciton polarons with interlayer interactions. Here, through magneto-optical reflectance contrast measurements, we experimentally investigate interlayer Fermi polarons for 2s excitons in WSe2/graphene heterostructures, where the excited exciton states (2s) in the WSe2 layer are dressed by free charge carriers of the adjacent graphene layer in the Landau quantization regime. First, such a system enables an optical detection of integer and fractional quantum Hall states (e.g., ν = ±1/3, ±2/3) of monolayer graphene. Furthermore, we observe that the 2s state evolves into two distinct branches, denoted as attractive and repulsive polarons, when graphene is doped out of the incompressible quantum Hall gaps. Our work paves the way for the understanding of the excited composite quasiparticles and Bose-Fermi mixtures.

A neural network potential energy surface of the Li3 system and quantum dynamics studies for the 7Li + 6Li2 → 6Li7Li + 6Li reaction.

A high-precision global potential energy surface (PES) of the Li3 system is constructed based on high-level ab initio calculations, and the root-mean-square error is 5.54 cm-1. The short-range of the PES is fitted by the fundamental invariant neural network (FI-NN) method, while the long-range uses a function with an accurate asymptotic potential energy form, and the two regions are connected by a switching function. Based on the new PES, the statistical quantum-mechanical (SQM) and the time-dependent wave packet (TDWP) methods are used to study the dynamics of 7Li + 6Li2 (v = 0, j = 0) → 6Li7Li + 6Li reactions in the low collision energy region (10-11 to 10-3 cm-1) and the high collision energy region (8 to 800 cm-1), respectively. In the high collision energy region, the calculation results of the SQM method and the TDWP method are inconsistent, indicating that the reaction dynamics does not follow the statistical behavior in the high collision energy region. In addition, we found that the Coriolis coupling effect plays an important role in this reaction. The symmetric forward-backward scattering in the total DCS indicates that the reaction follows the complex-forming reaction mechanism.

Regulating and controlling the stepwise ESDPT channel of BP(OH)2DCEt2 using the strategy of solvent polarity and external electric field.

Excited state double proton transfer (ESDPT) has attracted great scientific interest because of its excellent luminescent properties. However, the complex process of ESDPT has plagued theoretical and experimental scientists for a long time and has become a hot issue. In this work, the ESDPT process of 2,2'-bipyridine-3,3'-diol-5,5'-dicarboxylic acid ethyl ester (BP(OH)2DCEt2) is systematically studied and the regulation of the ESDPT process is further realized. The potential energy curves indicate that BP(OH)2DCEt2 shows the characteristics of stepwise ESDPT in different polar solvents. The increase in solvent polarity will be beneficial to the stepwise ESDPT reaction. Regrettably, it is not possible to distinguish the specific stepwise transfer path of the BP(OH)2DCEt2 molecule due to the symmetry of the potential energy surface along the diagonal. On this basis, we proposed a method to control and regulate the stepwise ESDPT path using an external electric field. The results show that the increase of external electric field intensity is favorable to stepwise ESDPT. It is interesting to note that applying an external electric field in a specific direction will effectively distinguish stepwise ESDPT reaction paths. Therefore, this work not only helps to understand the mechanism of ESDPT, but also contributes to regulation and design of new luminescent materials with excellent luminescent properties.

Quantum Dynamics Studies of the Li + Na2 (V = 0, j = 0) → Na + NaLi Reaction on a New Neural Network Potential Energy Surface.

The ultracold reaction offers a unique opportunity to elucidate the intricate microscopic mechanism of chemical reactions, and the Na2Li system serves as a pivotal reaction system in the investigation of ultracold reactions. In this work, a high-precision potential energy surface (PES) of the Na2Li system is constructed based on high-level ab initio energy points and the neural network (NN) method, and a proper asymptotic functional form is adopted for the long-range interaction, which is suitable for the study of cold or ultracold collisions. Based on the new NN PES, the dynamics of the Li + Na2 (v = 0, j = 0) → Na + NaLi reaction are studied in the collision energy range of 10-7 to 80 cm-1. In the high collision energy range of 8 to 80 cm-1, the dynamics of the reaction is studied using the time-dependent wave packet method and the statistical quantum mechanical (SQM) method. Comparing the results of the two methods, it is found that the SQM method provides a rough description of the product ro-vibrational state distribution but overestimates the integral cross-section values. With the decrease of collision energy, the reaction differential cross section gradually changes from forward-backward symmetric scattering to predominant forward scattering. In the low collision energy range from 10-7 to 8 cm-1, the SQM method is used to study the reaction dynamics, and the rate constant in the Wigner threshold region is estimated to be 2.87 × 10-10 cm3/s.

Gate-Tunable Multiband Transport in ZrTe5 Thin Devices.

Interest in ZrTe5 has been reinvigorated in recent years owing to its potential for hosting versatile topological electronic states and intriguing experimental discoveries. However, the mechanism of many of its unusual transport behaviors remains controversial: for example, the characteristic peak in the temperature-dependent resistivity and the anomalous Hall effect. Here, through employing a clean dry-transfer fabrication method in an inert environment, we successfully obtain high-quality ZrTe5 thin devices that exhibit clear dual-gate tunability and ambipolar field effects. Such devices allow us to systematically study the resistance peak as well as the Hall effect at various doping densities and temperatures, revealing the contribution from electron-hole asymmetry and multiple-carrier transport. By comparing with theoretical calculations, we suggest a simplified semiclassical two-band model to explain the experimental observations. Our work helps to resolve the longstanding puzzles on ZrTe5 and could potentially pave the way for realizing novel topological states in the two-dimensional limit.

Pygo-F773W Mutation Reveals Novel Functions beyond Wnt Signaling in Drosophila.

Pygopus (Pygo) has been identified as a specific nuclear co-activator of the canonical Wingless (Wg)/Wnt signaling pathway in Drosophila melanogaster. Pygo proteins consist of two conserved domains: an N-terminal homologous domain (NHD) and a C-terminal plant homologous domain (PHD). The PHD's ability to bind to di- and trimethylated lysine 4 of histone H3 (H3K4me2/3) appears to be independent of Wnt signaling. There is ongoing debate regarding the significance of Pygo's histone-binding capacity. Drosophila Pygo orthologs have a tryptophan (W) > phenylalanine (F) substitution in their histone pocket-divider compared to vertebrates, leading to reduced histone affinity. In this research, we utilized CRISPR/Cas9 technology to introduce the Pygo-F773W point mutation in Drosophila, successfully establishing a viable homozygous Pygo mutant line for the first time. Adult mutant flies displayed noticeable abnormalities in reproduction, locomotion, heart function, and lifespan. RNA-seq and cluster analysis indicated that the mutation primarily affected pathways related to immunity, metabolism, and posttranslational modification in adult flies rather than the Wnt signaling pathway. Additionally, a reduction in H3K9 acetylation levels during the embryonic stage was observed in the mutant strains. These findings support the notion that Pygo plays a wider role in chromatin remodeling, with its involvement in Wnt signaling representing only a specific aspect of its chromatin-related functions.

Use of 3D printing models for donor tooth extraction in autotransplantation cases.

OBJECTIVE: To clarify whether the 3D printing model has auxiliary functions in toto extraction of donor tooth in autotransplantation cases. METHODS: Two hundred and sixty patients who would have operation of ATT were divided into two groups. In group 1, determination of the tooth extraction in toto was predicted only according to the clinical and imaging examination. In group 2, the prediction was performed according to the clinical and imaging examination as well as the 3D model of donor tooth pre-extraction. A prespctive clinical study was designed on intra-group comparison between the predicted and actual donor teeth situation when extraction in cases of ATT. The consistent rate for the predicted results and the actual results were compared with the two groups. RESULTS: A remarkable difference was observed between the predicted results and the actual results of tooth positions and root numbers in group without model (p < 0,05). The consistency rate of the model group (94.62%) was significantly higher than that of non 3D model group (86.15%) (p = 0.034). CONCLUSION: The 3D printing model for the donor tooth is helpful for dentists to predict the accuracy of toto extraction of donor teeth in autotransplantation cases.

Microenvironment Modulation of Metal-Organic Frameworks (MOFs) for Coordination Olefin Oligomerization and (co)Polymerization.

The majority of commercial polyolefins are produced by coordination polymerization using early or late transition metal catalysts. Molecular catalysts containing these transition metals (Ti, Zr, Cr, Ni, and Fe, etc.) are loaded on supports for controlled polymerization behavior and polymer morphology in slurry or gas phase processes. Within the last few years, metal-organic frameworks (MOFs), a class of unique porous crystalline materials constructed from metal ions/clusters and organic ligands, have been designed and utilized as excellent supports for heterogeneous polymerization catalysis whose high density and uniform distribution of active sites would benefit the modulations of molecular weight distributions of high-performance olefin oligomers and (co)polymers. Impressive efforts have been made to modulate the microenvironment surrounding the active centers at the atomic level for improved activities of MOFs-based catalysts and controlled selectivity of olefin insertion. This review aims to draw a comprehensive picture of MOFs for coordination olefin oligomerization and (co)polymerization in the past decades with respect to different transition metal active centers, various incorporation sites, and finally microenvironment modulation. In consideration of more efforts are needed to overcome challenges for further industrial and commercial application, a brief outlook is provided.

The RNA helicase DDX5 promotes viral infection via regulating N6-methyladenosine levels on the DHX58 and NFκB transcripts to dampen antiviral innate immunity.

Multi-functional DEAD-box helicase 5 (DDX5), which is important in transcriptional regulation, is hijacked by diverse viruses to facilitate viral replication. However, its regulatory effect in antiviral innate immunity remains unclear. We found that DDX5 interacts with the N6-methyladenosine (m6A) writer METTL3 to regulate methylation of mRNA through affecting the m6A writer METTL3-METTL14 heterodimer complex. Meanwhile, DDX5 promoted the m6A modification and nuclear export of transcripts DHX58, p65, and IKKγ by binding conserved UGCUGCAG element in innate response after viral infection. Stable IKKγ and p65 transcripts underwent YTHDF2-dependent mRNA decay, whereas DHX58 translation was promoted, resulting in inhibited antiviral innate response by DDX5 via blocking the p65 pathway and activating the DHX58-TBK1 pathway after infection with RNA virus. Furthermore, we found that DDX5 suppresses antiviral innate immunity in vivo. Our findings reveal that DDX5 serves as a negative regulator of innate immunity by promoting RNA methylation of antiviral transcripts and consequently facilitating viral propagation.

Novel biphasic mechanism of the canonical Wnt signalling component PYGO2 promotes cardiomyocyte differentiation from hUC-MSCs.

Human umbilical cord-derived mesenchymal stem cells (hUC-MSCs) are used to regenerate the myocardium during cardiac repair after myocardial infarction. However, the regulatory mechanism underlying their ability to form mesodermal cells and differentiate into cardiomyocytes remains unclear. Here, we established a human-derived MSCs line isolated from healthy umbilical cords and established a cell model of the natural state to examine the differentiation of hUC-MSCs into cardiomyocytes. Quantitative RT-PCR, western blotting, immunofluorescence, flow cytometry, RNA Seq, and inhibitors of canonical Wnt signalling were used to detect the germ-layer markers T and MIXL1; the markers of cardiac progenitor cells MESP1, GATA4, and NKX2.5 and the cardiomyocyte-marker cTnT to identify the molecular mechanism associated with PYGO2, a key component of the canonical Wnt signalling pathway that regulates the formation of cardiomyocyte-like cells. We demonstrated that PYGO2 promotes the formation of mesodermal-like cells and their differentiation into cardiomyocytes through the hUC-MSC-dependent canonical Wnt signalling by promoting the early-stage entry of ß-catenin into the nucleus. Surprisingly, PYGO2 did not alter the expression of the canonical-Wnt, NOTCH, or BMP signalling pathways during the middle-late stages. In contrast, PI3K-Akt signalling promoted hUC-MSCs formation and their differentiation into cardiomyocyte-like cells. To the best of our knowledge, this is the first study to demonstrate that PYGO2 uses a biphasic mechanism to promote cardiomyocyte formation from hUC-MSCs.

H3K36 demethylase JMJ710 negatively regulates drought tolerance by suppressing MYB48-1 expression in rice.

The homeostasis of histone methylation is maintained by histone methyltransferases and demethylases, which are important for the regulation of gene expression. Here, we report a histone demethylase from rice (Oryza sativa), Jumonji C domain-containing protein (JMJ710), which belongs to the JMJD6 group and plays an important role in the response to drought stress. Overexpression of JMJ710 causes a drought-sensitive phenotype, while RNAi and clustered regularly interspaced short palindromic repeats (CRISPR)-knockout mutant lines show drought tolerance. In vitro and in vivo assays showed that JMJ710 is a histone demethylase. It targets to MYB TRANSCRIPTION FACTOR 48 (MYB48-1) chromatin, demethylates H3K36me2, and negatively regulates the expression of MYB48-1, a positive regulator of drought tolerance. Under drought stress, JMJ710 is downregulated and the expression of MYB48-1 increases, and the subsequent activation of its downstream drought-responsive genes leads to drought tolerance. This research reports a negative regulator of drought stress-responsive genes, JMJ710, that ensures that the drought tolerance mechanism is not mis-activated under normal conditions but allows quick activation upon drought stress.

Recombinase-mediated gene stacking in cotton.

Site-specific gene stacking could reduce the number of segregating loci and expedite the introgression of transgenes from experimental lines to field lines. Recombinase-mediated site-specific gene stacking provides a flexible and efficient solution, but this approach requires a recombinase recognition site in the genome. Here, we describe several cotton (Gossypium hirsutum cv. Coker 312) target lines suitable for Mycobacteriophage Bxb1 recombinase-mediated gene stacking. Obtained through the empirical screening of random insertion events, each of these target lines contains a single intact copy of the target construct with precise sequences of RS2, lox, and attP sites that is not inserted within or close to a known gene or near a centromere and shows good expression of the reporter gene gfp. Gene stacking was tested with insertion of different combinations of three candidate genes for resistance to verticillium wilt into three cotton target lines: CTS1, CTS3, and CTS4. Nine site-specific integration events were recovered from 95 independently transformed embryogenic calluses. Southern and DNA sequence analyses of regenerated plants confirmed precise site-specific integration, and resistance to verticillium wilt was observed for plant CTS1i3, which has a single precise copy of site-specifically integrated DNA. These cotton target lines can serve as foundation lines for recombinase-mediated gene stacking to facilitate precise DNA integration and introgression to field cultivars.

Rapid single-molecule digital detection of protein biomarkers for continuous monitoring of systemic immune disorders.

Digital protein assays have great potential to advance immunodiagnostics because of their single-molecule sensitivity, high precision, and robust measurements. However, translating digital protein assays to acute clinical care has been challenging because it requires deployment of these assays with a rapid turnaround. Herein, we present a technology platform for ultrafast digital protein biomarker detection by using single-molecule counting of immune-complex formation events at an early, pre-equilibrium state. This method, which we term "pre-equilibrium digital enzyme-linked immunosorbent assay" (PEdELISA), can quantify a multiplexed panel of protein biomarkers in 10 µL of serum within an unprecedented assay incubation time of 15 to 300 seconds over a 104 dynamic range. PEdELISA allowed us to perform rapid monitoring of protein biomarkers in patients manifesting post-chimeric antigen receptor T-cell therapy cytokine release syndrome, with ∼30-minute sample-to-answer time and a sub-picograms per mL limit of detection. The rapid, sensitive, and low-input volume biomarker quantification enabled by PEdELISA is broadly applicable to timely monitoring of acute disease, potentially enabling more personalized treatment.

Enhanced measurement of tiny rotational angles using conjugate orbital angular momentum modes.

We report the enhanced experimental measurement of tiny rotational angles using two conjugate OAM modes upon rotation of a Dove prism. The two conjugate OAM modes interfere in a petal-like pattern and the orientation of the pattern depends on the phase difference between the two modes. We propose an accurate method of digital image processing to measure the tiny rotational angles of the Dove prism. In the presence of an imperfect pattern and light path, the measurement precision was enhanced by a factor of l. This scheme has potential applications in high-precision sensing and monitoring of tiny rotation angles.

RNA helicase DExD/H-box 5 modulates intestinal microbiota in mice.

The RNA helicase DExD/H-box (DDX) family of proteins plays a central role in host cellular RNA metabolism, including mRNA regulation, microRNA biogenesis, and ribosomal processing. DDX5, also known as p68, promotes viral replication and tumorigenesis. However, there have been no studies on the regulation of the intestinal microbiota by DDX family proteins. We constructed DDX5 knockout mice (Ddx5+/-) using CRISPR/CAS9 technology. Subsequently, DDX5 knockout mice were analyzed for PCR products, mRNA levels, protein expression, immunohistochemistry, and histopathological lesions. Fecal (n = 12) and ileum (n = 12) samples were collected from the Ddx5+/- and wild-type (Ddx5+/+) mice. The diversity, richness, and structural separation of the intestinal microbiota of the Ddx5+/- and Ddx5+/+ mice were determined by 16S rRNA sequencing and analysis. Ddx5+/- mice were successfully established, and the ileum had normal morphology, a clear layer of tissue structures, and neatly arranged cupped cells. DDX5 knockout mice did not exhibit adverse effects on the ileal tissue. Microbial diversity and abundance were not significantly different, but the microbial structure of the intestinal microbiota was clustered separately between Ddx5+/+ and Ddx5+/- mice. Furthermore, we found that the relative abundance of Akkermansia and Clostridium_sensu_stricto_1 in the Ddx5+/- mice was significantly lower than in the Ddx5+/+ mice. These analyses indicated specific interactions between the intestinal microbiota and DDX5 protein. Our results indicate that DDX5 has a significant effect on the composition of the intestinal microbiota in mice, suggesting its potential as a promising novel target for the treatment of inflammation and tumorigenesis in the intestine.

Self-sampled specimens demonstrate comparable accuracy and consistency to clinician-sampled specimens for HPV detection among men who have sex with men in China.

OBJECTIVES: Despite a high risk of human papillomavirus (HPV) infection among men who have sex with men (MSM), few have ever tested. This study aimed to evaluate the feasibility and accuracy of HPV self-sampling among Chinese MSM, with the purpose of measuring the feasibility of self-sampling as an alternative in HPV testing scenarios. METHODS: Eligible participants were those who were assigned male at birth, aged 18 or above, had sex with men in the past year and had never gotten HPV vaccine. Participants followed the instructions to self-sample and were also clinician-sampled from the same anatomical sites (oral fluid, penis and rectum) in both approaches. All specimens were processed using multiplex PCR assay. The reference standard of an individual with a true positive for HPV is determined via PCR test, regardless of sampling methods. Sensitivity and specificity were calculated for each approach independently and kappa test was used to assess the consistency between the two approaches. RESULTS: Overall, 211 MSM were recruited at the local clinic from April to October 2020 in Zhuhai, China. The mean age was 31 years old. Only 3% of the participants sought help from healthcare providers during self-sampling. The prevalence of HPV was 49% (103 of 211). Clinician sampling detected 91 of 103 MSM infected with HPV, with a sensitivity of 88.3% (95% CI 80.2 to 93.6) and a specificity of 100.0% (95% CI 95.7 to 100.0). Self-sampling detected 81 of 103 MSM infected with HPV, with a sensitivity of 78.6% (95% CI 69.2 to 85.9) and a specificity of 100.0% (95% CI 95.7 to 100.0). The level of agreement was moderate between clinician sampling and self-sampling (k=0.67). CONCLUSIONS: Self-sampled HPV testing demonstrated comparable accuracy and consistency to clinician sampling among MSM in China. It holds the potential to complement sexual health services especially among key populations.

A neutralizing monoclonal antibody-based blocking ELISA to detect bovine herpesvirus 1 and vaccination efficacy.

Infections caused by bovine herpesvirus 1 (BoHV-1) remain a serious global issue to the health and welfare of the bovine industry. Monitoring of neutralizing antibodies is essential not only for epidemic diagnosis, but also to assess vaccination efficacy. In this study, we generated a neutralizing monoclonal antibody, termed as 3F8, targeting glycoprotein D (gD) of BoHV-1. This monoclonal antibody could neutralize BoHV-1 with a 50% inhibitory concentration (IC50) of 37.82 ng/mL. Furthermore, 3F8 could inhibit BoHV-1 infection and cell-to-cell spread at the prebinding stage. A blocking enzyme-linked immunosorbent assay (ELISA) for detecting neutralizing antibodies against BoHV-1 was then developed based on 3F8 and protein gD generated using a baculovirus expression system. The sensitivity and specificity of the test were estimated to be 94.59% and 93.42%, respectively. A significant correlation (R2 = 0.9583, p < 0.01) was observed between the results obtained with the blocking ELISA and a virus neutralization test, which suggested that the blocking ELISA could detect neutralizing antibodies against BoHV-1. A serological survey was carried out in the dairy farms in Beijing district using 3F8-based blocking ELISA to monitor the annual neutralization antibody against BoHV-1 during 2012-2020. It revealed that the dairy farms in Beijing were at high risk of BoHV-1 infection during 2012-2017 but were protected since 2018 upon implementation of an immunization program. Our results demonstrated that this assay is suitable for BoHV-1 surveillance and vaccination efficacy in cattle as a replacement for the virus neutralization test. KEY POINTS: • Prevention of BoHV-1 infection requires the monitoring of neutralizing antibodies. • A blocking ELISA for the neutralizing antibody was developed based on mAb 3F8 against BoHV-1 gD. • It can replace the labor-intensive and time-consuming viral neutralizing tests.

asb5a/asb5b Double Knockout Affects Zebrafish Cardiac Contractile Function.

Ankyrin repeat and suppression-of-cytokine-signaling box (Asb) proteins, a subset of ubiquitin ligase E3, include Asb5 with six ankyrin-repeat domains. Zebrafish harbor two asb5 gene isoforms, asb5a and asb5b. Currently, the effects of asb5 gene inactivation on zebrafish embryonic development and heart function are unknown. Using CRISPR/Cas9, we generated asb5a-knockout zebrafish, revealing no abnormal phenotypes at 48 h post-fertilization (hpf). In situ hybridization showed similar asb5a and asb5b expression patterns, indicating the functional redundancy of these isoforms. Morpholino interference was used to target asb5b in wild-type and asb5a-knockout zebrafish. Knocking down asb5b in the wild-type had no phenotypic impact, but simultaneous asb5b knockdown in asb5a-knockout homozygotes led to severe pericardial cavity enlargement and atrial dilation. RNA-seq and cluster analyses identified significantly enriched cardiac muscle contraction genes in the double-knockout at 48 hpf. Moreover, semi-automatic heartbeat analysis demonstrated significant changes in various heart function indicators. STRING database/Cytoscape analyses confirmed that 11 cardiac-contraction-related hub genes exhibited disrupted expression, with three modules containing these genes potentially regulating cardiac contractile function through calcium ion channels. This study reveals functional redundancy in asb5a and asb5b, with simultaneous knockout significantly impacting zebrafish early heart development and contraction, providing key insights into asb5's mechanism.