Biology subjects

Peng, X.

Publications and source records attributed to Peng, X..

At least 109 records · Page 6Linked to original sources

bioRxiv2023

RNPS1 stabilizes NAT10 protein to facilitate translation in cancer via tRNA ac4C modification

NAT10 is dysregulated and plays an essential role in various types of cancers. However, the exact machenism of how NAT10 regulates cancer progression remains debatable. In this report, we show that NAT10 affects tumorigeneis mainly based on its acetylation function on tRNA. In addition, we found NAT10 regulate the ac4C of tRNA in cancer via interaction with RNPS1, which in turn protect NAT10 from degradation by E3 ubiquitin ligase ZSWIM6. We developed TRMC-seq method to compreshensively profile tRNA ac4C sites and uncovered the presence of ac4C in a broader range of tRNA isoacceptors than previous studies. Multi-omics analysis identified AP-1 signaling pathway as a major downstream mediator of NAT10. Mechanistically, we found NAT10 is responsible for the translation efficiency genes which contain higher ac4C-tRNA codon. Importantly, our genetic mouse model validated our in vitro findings of NAT10 in cancer. Our study highlights a role of NAT10 in mediating tRNA ac4C to regulate the translation and tumorigenesis of cancer.

molecular biology↗

bioRxiv2023

ARPC5 deficiency leads to severe early onset systemic inflammation and early mortality

The seven subunit Arp2/3 complex drives the formation of branched actin networks that are essential for many cellular processes including cell migration. In humans, the ARPC5 subunit of the Arp2/3 complex is encoded by two paralogous genes (ARPC5 and ARPC5L), resulting in proteins with 67% identity. Through whole-exome sequencing, we identified a biallelic ARPC5 frameshift variant in a female child who presented with recurrent infections, multiple congenital anomalies, diarrhea, and thrombocytopenia, and suffered early demise from sepsis. Her consanguineous parents also had a previous child who died with similar clinical features. Using CRISPR/Cas9-mediated approaches, we demonstrate that loss of ARPC5 affects actin cytoskeleton organization and function, as well as chemokine-dependent cell migration in vitro. Homozygous Arpc5-/- mice do not survive past embryonic day 9 due to severe developmental defects, including loss of the second pharyngeal arch which contributes to craniofacial and heart development. Our results indicate that ARPC5 is important for both prenatal development and postnatal immune signaling, in a non-redundant manner with ARPC5L. Moreover, our observations add the ARPC5 locus to the list of genes that should be considered when patients present with syndromic early-onset immunodeficiency, particularly if recessive inheritance is suspected.

immunology↗

bioRxiv2023

To be or not to be an anti-CRISPR: AcrIII-1 and the importance of working with native biological systems

Viral members of the protein family DUF1874 have been reported to act as anti-CRISPR (acr) proteins that degrade cyclic tetra-adenylate (cA4), a nucleotide second messenger produced after the activation of several type III CRISPR-Cas systems in bacteria and archaea. Specifically, protein SIRV1 gp29 inhibits type III-A and type III-B CRISPR systems in plasmid-born assays in heterologous systems. In this work, we investigate the function of SIRV1 gp29 and its close homolog SIRV2 gp37 in a native biological context, i.e. in cultures infected by SIRV2. SIRV2 was selected instead of SIRV1 because the latter is not available any more from any laboratories. We demonstrate that gp37 has no anti-CRISPR activity during infection of Saccharolobus islandicus LAL14/1 with SIRV2, although it is able to protect SIRV2 from type III targeting when expressed from a plasmid. The inability of gp37 to act as an acr in the native, biological system is due to the protein expression timing: gp37 is a middle/late gene, thus unable to inhibit CRISPR-Cas targeting at the onset of infection. On the other hand, we find that while gp37 is a non-essential gene, it confers a mild replicative advantage to the virus. This advantage is mediated, in hosts with active CRISPR-Cas targeting, by the interaction between gp37 and host protein SiL_1451, which results in the inhibition of the lysine methyltransferase activity of SiL_1451, responsible for extensive methylation of surface lysines of two-thirds of the cellular proteins. Heterologous gene expression from a plasmid-borne non-native promoter has allowed the discovery and characterization of dozens of prokaryotic defense systems in recent years. Although this experimental strategy has several advantages, our study highlights the importance of validating relevant results under native conditions, and the limitations of extrapolating results obtained using heterologous systems.

microbiology↗

bioRxiv2022

DPYSL2/CRMP2 isoform B knockout in human iPSC-derived glutamatergic neurons confirms its role in mTOR signaling and neurodevelopmental disorders

DPYSL2/CRMP2 is a microtubule-stabilizing protein crucial for neurogenesis and associated with numerous psychiatric and neurodegenerative disorders. DPYSL2 has multiple RNA and protein isoforms, but few studies have differentiated between them or explored their individual functions. We previously demonstrated in HEK293 cells that a schizophrenia -associated variant in the DPYSL2 B isoform (DPYSL2-B) reduced the length of cellular projections, created a transcriptomic disturbance that captured schizophrenia etiology, and was acted upon by the mTOR pathway. In the present study, we follow up on these results by creating, to our knowledge, the first models of endogenous DPYSL2-B knockout in human induced pluripotent stem cells and excitatory glutamatergic neurons. We use CRISPR/Cas9 to specifically knock out DPYSL2-B and observe corresponding reduction of its RNA and protein. The average length of dendrites in knockout neurons was reduced up to 58% compared to controls. Transcriptome analysis reveals disruptions in pathways highly relevant to psychiatric disease including mTOR signaling, cytoskeletal dynamics, immune function, calcium signaling, and cholesterol biosynthesis. We also observed a significant enrichment of our differentially expressed genes in schizophrenia GWAS-associated loci. Our findings clarify the functions of the human DPYSL2-B isoform and confirm its involvement in molecular pathologies shared between many psychiatric diseases.

genetics↗

bioRxiv2022

Proliferative Exhausted CD8 T Cells Exacerbate Long-lasting Anti-tumor Effects in Human Papillomavirus Positive Head and Neck Squamous Cell Carcinoma

The survival prognosis of human papillomavirus (HPV)-positive and HPV-negative head and neck squamous cell carcinoma (HNSCC) is largely different, and little is known about the anti-tumor mechanism of tumor-infiltrated CD8+ exhausted T cells (Tex) in HNSCC. We performed cell level multi-omics sequencing on human HNSCC samples to decipher the multi-dimensional characteristics of Tex cells. A proliferative exhausted CD8+ T cell cluster (P-Tex) which was beneficial to survival outcomes of patients with HPV-positive HNSCC was identified. Interestingly, P-Tex cells expressed CDK4 genes as high as cancer cells, which could be simultaneously inhibited by CDK4 inhibitors and might be a potential reason for the ineffectiveness of CDK4 inhibitors in treating HPV-positive HNSCC. P-Tex cells could aggregate in the antigen presenting cell niches and activate certain signaling pathways. Together, our findings suggest a promising role for P-Tex cells in the prognosis of patients with HPV-positive HNSCC by providing modest but persistent anti-tumor effects. Graphic abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=105 SRC="FIGDIR/small/513993v1_ufig1.gif" ALT="Figure 1"> View larger version (39K): org.highwire.dtl.DTLVardef@1f8810forg.highwire.dtl.DTLVardef@1e11d84org.highwire.dtl.DTLVardef@58fb6aorg.highwire.dtl.DTLVardef@1e20a13_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2022

Characterization of Three Variants of SARS-CoV-2 in vivo Shows Host-Dependent Pathogenicity in Hamsters

Animal models are used in preclinical trials to test vaccines, antivirals, monoclonal antibodies, and immunomodulatory drug therapies against SARS-CoV-2. However, these drugs often do not produce equivalent results in human clinical trials. Here, we show how different animal models infected with some of the most clinically relevant SARS-CoV-2 variants, WA1/2020, B.1.617.2/Delta, B.1.1.529/Omicron and BA5.2/Omicron, have independent outcomes. We show that in mice, B.1.617.2 is more pathogenic, followed by WA1, while B.1.1.529 showed an absence of clinical signs. Only B.1.1.529 was able to infect C57BL/6J mice, which lack the human ACE2 receptor. B.1.1.529-infected ACE2 mice had different T cell profiles compared to infected K18-hACE2 mice, while viral shedding profiles and viral titers in lungs were similar between the ACE2 and the C57BL/6J mice. These data suggest B.1.1.529 virus adaptation to a new host and shows that asymptomatic carriers can accumulate and shed virus. Next, we show how B.1.617.2, WA1 and BA5.2/Omicron have similar viral replication kinetics, pathogenicity, and viral shedding profiles in hamsters, demonstrating that the increased pathogenicity of B.1.617.2 observed in mice is host-dependent. Overall, these findings suggest that small animal models are useful to parallel human clinical data, but the experimental design places an important role in interpreting the data. IMPORTANCEThere is a need to investigate SARS-CoV-2 variants phenotypes in different animal models due to the lack of reproducible outcomes when translating experiments to the human population. Our findings highlight the correlation of clinically relevant SARS-CoV-2 variants in animal models with human infections. Experimental design and understanding of correct animal models are essential to interpreting data to develop antivirals, vaccines, and other therapeutic compounds against COVID-19.

microbiology↗

bioRxiv2022

Mechanobiology of Fibroblast Activation in Skin Grafting

Mechanical stretching of living tissues can activate long-lived changes in tissue cells such as fibroblasts, increasing their contractility and initiating phenotypic transformations. Increased mechanical stimulus typically leads to monotonically increasing activation of fibroblasts cultured in 2D, but activation levels are difficult to predict for cells in 3D fibrous tissues, leading to variable outcomes in procedures such as skin grafting. Here we report that the source of this variation is cell-extracellular matrix (ECM) interactions and their variation with the duration and magnitude of applied stretch, and present a model that can predict the degree to which stretch will either increase or decrease long-term activation levels of fibroblasts cultured within a stretched, three-dimensional collagen matrix. Combining experimental and mathematical approaches across multiple scales, we show that the viscoplasticity of the ECM regulates this nonmonotonic, long-term cell activation. Results demonstrate that feedback between cell and ECM determines how cells retain memory of mechanical stretch.

biophysics↗

bioRxiv2022

Intrinsically disordered proteins SAID1/2 condensate on SERRATE/ARS2 for dual inhibition of miRNA biogenesis in Arabidopsis

Intrinsically disordered proteins (IDPs) SAID1/2 are hypothetic dentin sialophosphoprotein-like proteins, but their true functions are unknown. Here, we identified SAID1/2 as negative regulators of SERRATE(SE)/ARS2, a core factor in miRNA biogenesis complex (microprocessor). Loss-of-function double mutants of said1; said2 caused pleiotropic developmental defects and thousands of differentially-expressed genes that partially overlapped with those in se. said1; said2 also displayed increased assembly of microprocessor and elevated accumulation of miRNAs. Mechanistically, SAID1/2 promotes PRP4KA-mediated phosphorylation of SE, leading to its degradation in vivo. Unexpectedly, SAID1/2 have strong binding affinity to hairpin-structured pri-miRNAs and can sequester them from SE. Moreover, SAID1/2 directly inhibit pri-miRNA processing by microprocessor in vitro. Whereas SAID1/2 did not impact SE subcellular compartmentation, the proteins themselves exhibited liquid-liquid phase condensation that is nucleated on SE. Thus, we proposed that SAID1/2 reduce miRNA production through hijacking pri-miRNAs to prevent microprocessor activity while promoting SE phosphorylation and its destabilization in Arabidopsis.

plant biology↗

bioRxiv2022

Loss of H3.1K27me1 in Arabidopsis confers resistance to Geminivirus by sequestering DNA repair proteins onto rDNA and defense-related genes

The H3 methyltransferases ATXR5 and ATXR6 deposit H3.1K27me1 to heterochromatin to prevent genomic instability and transposon reactivation. Here, we report that atxr5 atxr6 mutants displayed robust resistance to Geminivirus. The viral resistance correlated with activation of DNA repair pathways, but not with transposon reactivation or heterochromatin amplification. We identified RAD51 and RPA1A as partners of virus-encoded Rep protein. The two DNA repair proteins showed increased binding to heterochromatic regions and defense-related genes in atxr5 atxr6 vs wild type plants. Consequently, the proteins had reduced interactions to viral DNA in the mutant, thus hampering viral replication. Additionally, RAD51 recruitment to the host genome arose via BRCA1, HOP2 and CYCB1, and this recruitment was essential for viral resistance in atxr5 atxr6. Thus, Geminiviruses adapt to healthy plants by hijacking its DNA repairing pathways for replication, but the host could retain DNA repairing proteins via sacrificing its genome stability to suppress viral infection.

genetics↗

bioRxiv2022

ApoE4 causes severe COVID-19 outcomes via downregulation of ACE2

The coronavirus disease 2019 (COVID-19) pandemic is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2); host cell entry by this virus relies on the interaction between the receptor-binding domain (RBD) of its spike glycoprotein and the angiotensin-converting enzyme 2 (ACE2) receptor on cell membranes. In addition to serving as a receptor for SARS-CoV-2, ACE2 was originally discovered as a protective factor in the renin-angiotensin system (RAS) that catalyses the degradation of angiotensin II (Ang II) to Ang 1-7, which is involved in multiple organ pathology. Recent genetic and clinical studies reported that ApoE4 expression is associated with increased susceptibility to SARS-CoV-2 infection and the development of severe COVID-19, but the underlying mechanism is currently unclear. In the present study, by using immunofluorescence staining, molecular dynamics simulations, proximity ligation assay (PLA) and coimmunoprecipitation (Co-IP) combined with a biolayer interferometry (BLI) assay, we found that ApoE interacts with both the spike protein and ACE2 but does not show obvious isoform-dependent binding effects. These data suggest that ApoE4 increases SARS-CoV-2 infectivity in a manner that may not depend on differential interactions with the spike protein or ACE2. Importantly, further immunoblotting and immunofluorescence staining results showed that ApoE4 significantly downregulates ACE2 protein expression in vitro and in vivo and subsequently decreases the conversion of Ang II to Ang 1-7, which could worsen tissue lesions; these findings provide a possible explain by which ApoE4 exacerbates COVID-19 disease.

neuroscience↗

bioRxiv2022

Linking mesozooplankton and SAR11 bacteria in Oxygen Deficient Zones and the open ocean

The gravitational biological pump is not large enough to account for microbial heterotrophic activity in the mesopelagic ocean. Migrating zooplankton may be a key source of organic matter transport to depth. Here we show signatures of zooplankton in the suspended organic matter at the zooplankton vertical migration depth in the Eastern Tropical North Pacific Oxygen Deficient Zone (ETNP ODZ). We examine the mesozooplankton community in metagenomic depth profiles using the mitochondrial cytochrome c oxidase (COI) gene as a marker in the ETNP and Eastern Tropical South Pacific (ETSP) ODZs and at the oxic Hawaii Ocean Timeseries (HOT). Additionally, eukaryotic transcripts (polyA-selected) were examined for zooplankton in the ETNP. While zooplankton eDNA increased in the ODZ, zooplankton eRNA decreased in the ODZ, similar to previous net-based data, implying that eDNA is better preserved under anoxia. At all stations, Cnidaria, often missed in net-based data, contributed greatly to the zooplankton eDNA/eRNA. SAR11 abundance, determined from the single-copy core gene (rpoB), significantly correlated with zooplankton eDNA, with R2 values >0.8 at all stations. Strong correlations between SAR11 and zooplankton have not been previously reported, but are logical as SAR11 bacteria consume and zooplankton excrete simple dissolved organic compounds. SAR11 bacteria possessed genes to utilize urea and taurine in the ODZ, both compounds known to be excreted by zooplankton. In ODZs, SAR11 bacteria preferentially used the taurine degradation pathway leading to C and N assimilation, not the pathway for organic S assimilation, probably due to additional sources of organic S in ODZs.

microbiology↗

bioRxiv2022

A clade of RHH proteins ubiquitous in Sulfolobales and their viruses regulates cell cycle progression

Cell cycle regulation is crucial for all living organisms and is often targeted by viruses to facilitate their own propagation, yet cell cycle progression control is largely underexplored in archaea. In this work, we reveal a cell cycle regulator (aCcr1) carrying a ribbon-helix-helix (RHH) domain and ubiquitous in the crenarchaeal order Sulfolobales and their viruses. Overexpression of several aCcr1 members including gp21 of rudivirus SIRV2 and its host homolog SiL_0190 of S. islandicus LAL14/1 results in impairment of cell division, evidenced by growth retardation, cell enlargement and an increase in cellular DNA content. Additionally, both gp21 and SiL_0190 can bind to the motif AGTATTA conserved in the promoter of several genes involved in cell division, DNA replication and cellular metabolism thereby repressing or inducing their transcription. Our results suggest that aCcr1 silences cell division and drives progression to the S-phase in Sulfolobales, a function exploited by viruses to facilitate viral propagation.

microbiology↗

bioRxiv2022

Replidec - Use naive Bayes classifier to identify virus lifecycle from metagenomics data

MotivationViruses are the most abundant biological entities on earth. The majority of these entities are bacterial viruses or phages which specifically infect bacteria. Phages can use different replication strategies to invade their hosts including lytic, lysogenic, chronic cycle and pseudolysogeny. While the determination of the replication strategy used by phages is important to explore the phage-bacteria relationships in different ecosystems there are not many tools that can predict this in metagenomic data. In addition, most of the tools available can only predict lytic and lysogenic cycles. To address this issue, we have developed a new software called Replidec to identify three most common phage replication cycles (virulent, temperate, chronic) in viral sequences. ResultsReplidec uses Naive Bayes classifier combined with alignment-based methods to improve the prediction accuracy in metagenomic data. We test Replidec on viral genomes with known replication cycle and simulated metagenomic sequences. Replidec perform relatively good both in isolated genomes (F1 score: 92.29% {+/-} 0.81; mcc: 89.14% {+/-} 1.22) and simulated metagenomic sequences(F1 score: 87.55% {+/-} 2.12; mcc: 88.23% {+/-} 2.55). Moreover, Replidec can also accurately predict the replication cycle in small viral fragments([~]3000bp). In conclusion, Replidec can achieve the best performance in simulated metagenomic data compared to most prediction softwares including BACPHLIP.

bioinformatics↗

bioRxiv2022

Antigen-Specific Antibody Design and Optimization with Diffusion-Based Generative Models

Antibodies are immune system proteins that protect the host by binding to specific antigens such as viruses and bacteria. The binding between antibodies and antigens is mainly determined by the complementarity-determining regions (CDR) of the antibodies. In this work, we develop a deep generative model that jointly models sequences and structures of CDRs based on diffusion probabilistic models and equivariant neural networks. Our method is the first deep learning-based method that generates antibodies explicitly targeting specific antigen structures and is one of the earliest diffusion probabilistic models for protein structures. The model is a "Swiss Army Knife" capable of sequence-structure co-design, sequence design for given backbone structures, and antibody optimization. We conduct extensive experiments to evaluate the quality of both sequences and structures of designed antibodies. We find that our model could yield competitive results in binding affinity measured by biophysical energy functions and other protein design metrics.

bioinformatics↗

bioRxiv2022

Targeting Chromatin Effector Pygo2 to Enhance Immunotherapy in Prostate Cancer

The noninflamed microenvironment in prostate cancer represents a barrier to immunotherapy. Genetic alterations underlying cancer cell-intrinsic oncogenic signaling have emerging roles in shaping the immune landscape. Recently, we identified Pygopus 2 (PYGO2) as the driver oncogene for the amplicon at 1q21.3. Here, using transgenic mouse models of metastatic prostate adenocarcinoma, we found that Pygo2 deletion decelerated tumor progression, diminished metastases, and extended survival. Pygo2 loss augmented the infiltration of cytotoxic T lymphocytes (CTLs) and sensitized tumor cells to T cell killing. Mechanistically, Pygo2 orchestrated a p53/Sp1/Kit/Ido1 signaling network to foster a microenvironment hostile to CTLs. Genetic or pharmacological inhibition of Pygo2 enhanced the anti-tumor efficacy of immunotherapies using immune checkpoint blockade, adoptive cell transfer, or myeloid-derived suppressor cell inhibitors. In human prostate cancer samples, Pygo2 expression was inversely correlated with CD8+ T cells. Our results highlight a promising path to improving immunotherapy with targeted therapy for lethal prostate cancer.

cancer biology↗

bioRxiv2022

Accurate Estimation of Molecular Counts from Amplicon Sequence Data with Unique Molecular Identifiers

MotivationAmplicon sequencing is widely applied to explore heterogeneity and rare variants in genetic populations. Resolving true biological variants and quantifying their abundance is crucial for downstream analyses, but measured abundances are distorted by stochasticity and bias in amplification, plus errors during Polymerase Chain Reaction (PCR) and sequencing. One solution attaches Unique Molecular Identifiers (UMIs) to sample sequences before amplification eliminating amplification bias by clustering reads on UMI and counting clusters to quantify abundance. While modern methods improve over naive clustering by UMI identity, most do not account for UMI reuse, or collision, and they do not adequately model PCR and sequencing errors in the UMIs and sample sequences. ResultsWe introduce Deduplication and accurate Abundance estimation with UMIs (DAUMI), a probabilistic framework to detect true biological sequences and accurately estimate their deduplicated abundance from amplicon sequence data. DAUMI recognizes UMI collision, even on highly similar sequences, and detects and corrects most PCR and sequencing errors in the UMI and sampled sequences. DAUMI performs better on simulated and real data compared to other UMI-aware clustering methods. AvailabilitySource code is available at https://github.com/xiyupeng/AmpliCI-UMI.

bioinformatics↗

bioRxiv2022

Suppression of tumor/host intrinsic CMTM6 drives anti-tumor cytotoxicity in a PD-L1 independent manner

CKLF-like MARVEL transmembrane domain-containing 6 (CMTM6) has been identified as a regulator of membranal programmed death ligand 1 (PD-L1) stability and a factor associated with malignancy progression, but the effects and mechanisms of CMTM6 on tumor growth, as well as its potential for therapy, are still largely unknown. Here, we show that tumor CMTM6 increased with progression in both clinical patients and mice. Ablation of CMTM6 resulted in significant retardation of human and murine tumor growth dependent on T-lymphocyte immunity. Tumor CMTM6 suppression broke resistance to immune checkpoint inhibitors and remodeled the tumor immune microenvironment, as specific antitumor cytotoxicity was enhanced and contributed primarily to tumor inhibition. Further, without the PD-1/PD-L1 axis, CMTM6 suppression still significantly dampened tumor growth dependent on cytotoxic cells. Notably, we identified that CMTM6 was widely expressed on immune cells. T-cell CMTM6 increased with sustained immune activation and intratumoral immune exhaustion and affected the T-cell-intrinsic PD-L1 levels. Host CMTM6 knockout significantly restrained tumor growth dependent on CD8+ T-cells, and similarly, not entirely dependent on PD-L1. Thus, we developed and evaluated the antitumor efficacy of CMTM6-targeting adeno-associated virus (AAV), which effectively mobilized antitumor immunity and could be combined with various antitumor drugs. Our findings reveal that both tumor and host CMTM6 are deeply involved in tumor immunity with or without the PD-1/PD-L1 axis and that gene therapy targeting CMTM6 is a promising strategy for cancer immunotherapy. One Sentence SummaryEven in the absence of the PD-1/PD-L1 axis, tumor or host CMTM6 deficiency can mediate cytotoxicity-dependent anti-tumor immune responses, allowing CMTM6 to be a novel target for scAAV-mediated oncoimmunology gene therapy and combination treatment.

immunology↗

bioRxiv2022

Hinokitiol alters Gene Expression in Aspergillus fumigatus, protects against fungal keratitis by Reducing Fungal Load, LOX-1, Proinflammatory cytokines and Neutrophil Infiltration

Hinokitiol (HK), a tropolone-related compound found in cupressaceous plants, has antifungal and anti-inflammatory properties. But its roles in Aspergillus fumigatus ( A.F. ) keratitis remains unknown. This study examined the antifungal activity of HK against A.F. and investigated its possible mechanisms at the ultrastructural and transcriptional levels. We tested the minimum inhibitory concentration (MIC) of HK against A.F.. Our data showed that the MIC50 was 2 g/ml and MIC90 was 8 g/ml, and HK could significantly inhibit the spore germination and mycelial growth of A.F.. Scanning electron microscopy (SEM) and transmission electron microscopy (TEM) observations showed that HK induced significant changes in hyphal morphology and microstructure, including cell membrane rupture and intracellular structural disorder. Adhesion assay and biofilm assay experiments showed that HK reduced the adhesion of spores to human corneal epithelial cells (HCECs) and inhibited the formation of biofilms. The genetic changes of A.F. after HK treatment were analyzed by RNA sequencing, and a total of 2487 differentially expressed genes (DEGs) were identified, including the down-regulated DEGs related to carbohydrate and various amino acid metabolism, ribosome biogenesis, asexual reproduction, and up-regulated DEGs related to steroid biosynthesis through GO and KEGG enrichment analysis. RNA-seq results suggested that HK may play an antifungal effect by destroying the structural integrity of its wall and membrane, inhibiting protein biosynthesis and the growth and reproduction of A.F..10 g/ml HK, which had no effect on proliferation and migration of HCECs, was used in the treatment of murine fungal keratitis (FK). We found that HK eyedrop treatment could reduce the severity of FK by reducing corneal fungal burden, neutrophil infiltration, and the expression of LOX-1 and pro-inflammatory factors. HK is expected to be a safe and effective new drug for FK treatment. Author summaryFK is a serious common blind-causing eye disease. The current clinical antifungal drugs have the disadvantages of low bioavailability and high corneal toxicity, which greatly limit the clinical effectiveness. Therefore, there is an urgent need for a new, safe and effective treatment for FK. HK had shown great antifungal capacity against A.F., amazingly well anti-inflammatory effect and real improvement in murine FK in our research. These findings provide a theoretical basis for the application of HK as an antifungal drug and reveal the potential of HK to be applied in the clinical treatment of FK.

microbiology↗