A1 Domain Research Articles

Mucosal LTA1 Elicits Lung Th17 cells with reduced plasticity towards Th1 cells and resistance to FK506.

Abstract Objective: Based on our previous work, we identified a vaccine composed of the A1 domain of heat-labile toxin from E. coli (LTA1) adjuvant and Outer membrane protein (Omp) X from K2 strain of Klebsiella pneumoniae (K. pneumoniae), which can elicit serotype independent protection against K1 strain by generating lung Th1 and Th17 cells. To better understand the characteristics of these Th17 cells, we investigated the TCR repertoire, plasticity, and resistance to FK506. Methods: Naïve or immunized lung cells were collected and stained with Anti-Mouse TCR Vβ Screening Panel for FACS. Naïve CD4+ cells were isolated from spleens and cultured under Th17 differentiation conditions in vitro. Immunized lung Th17 cells were purified. To assess plasticity towards Th1, cells were cultured w/wo IL-12 and analyzed by FACS. To assess resistance to calcineurin inhibitors, the naïve/immunized CD4+ cells were treated w/wo FK506 both in vivo and ex vivo, analyzed by FACS. Results and Conclusions: The LTA1-OmpX elicited CD4+ cells were oligoclonal with skewing towards TCRVβ14+ cells. After incubation with IL-12, 57.73 ± 3.9% of the in vitro differentiated Th17 cells showed plasticity towards Th1 cells whereas in immunized lung Th17 cells only showed 6.25 ± 1.6%. Lung Th17 cells also showed substantially reduced STAT4 phosphorylation and surface IL-12rβ2 expression compared to in vitro Th17 cells. Furthermore, lung Th17 cell IL-17A expression was resistant to FK506 treatment both in vivo and ex vivo, which potentially indicated that this vaccine strategy may confer protection in the setting of transplant immunosuppression.

Clinical spectrum and genetic variations of LMNA-related muscular dystrophies in a large cohort of Chinese patients

BackgroundLMNA-related muscular dystrophy is caused by mutations in LMNA gene. We aimed to identify genetic variations and clinical features in a large cohort of Chinese patients with LMNA mutations in...

Activation of von Willebrand factor via mechanical unfolding of its discontinuous autoinhibitory module

Von Willebrand factor (VWF) activates in response to shear flow to initiate hemostasis, while aberrant activation could lead to thrombosis. Above a critical shear force, the A1 domain of VWF becomes activated and captures platelets via the GPIb-IX complex. Here we show that the shear-responsive element controlling VWF activation resides in the discontinuous autoinhibitory module (AIM) flanking A1. Application of tensile force in a single-molecule setting induces cooperative unfolding of the AIM to expose A1. The AIM-unfolding force is lowered by truncating either N- or C-terminal AIM region, type 2B VWD mutations, or binding of a ristocetin-mimicking monoclonal antibody, all of which could activate A1. Furthermore, the AIM is mechanically stabilized by the nanobody that comprises caplacizumab, the only FDA-approved anti-thrombotic drug to-date that targets VWF. Thus, the AIM is a mechano-regulator of VWF activity. Its conformational dynamics may define the extent of VWF autoinhibition and subsequent activation under force.

Quantification of von Willebrand factor and ADAMTS-13 after traumatic injury: a pilot study

BackgroundVon Willebrand factor (VWF) is an acute phase reactant synthesized in the megakaryocytes and endothelial cells. VWF forms ultra-large multimers (ULVWF) which are cleaved by the metalloprotease ADAMTS-13, preventing spontaneous...

Sialylation on O-linked glycans protects von Willebrand factor from macrophage galactose lectin-mediated clearance.

Terminal sialylation determines the plasma half-life of von Willebrand factor (VWF). A role for macrophage galactose lectin (MGL) in regulating hyposialylated VWF clearance has recently been proposed. In this study, we showed that MGL influences physiological plasma VWF clearance. MGL inhibition was associated with a significantly extended mean residence time and 3-fold increase in endogenous plasma VWF antigen levels (P<0.05). Using a series of VWF truncations, we further demonstrated that the A1 domain of VWF is predominantly responsible for enabling the MGL interaction. Binding of both full-length and VWF-A1-A2-A3 to MGL was significantly enhanced in the presence of ristocetin (P<0.05), suggesting that the MGL-binding site in A1 is not fully accessible in globular VWF. Additional studies using different VWF glycoforms demonstrated that VWF O-linked glycans, clustered at either end of the A1 domain, play a key role in protecting VWF against MGLmediated clearance. Reduced sialylation has been associated with pathological, increased clearance of VWF in patients with von Willebrand disease. Herein, we demonstrate that specific loss of α2-3 linked sialylation from O-glycans results in markedly increased MGL-binding in vitro, and markedly enhanced MGL-mediated clearance of VWF in vivo. Our data further show that the asialoglycoprotein receptor (ASGPR) does not have a significant role in mediating the increased clearance of VWF following loss of O-sialylation. Conversely however, we observed that loss of N-linked sialylation from VWF drives enhanced circulatory clearance predominantly via the ASGPR. Collectively, our data support the hypothesis that in addition to regulating physiological VWF clearance, the MGL receptor works in tandem with ASGPR to modulate enhanced clearance of aberrantly sialylated VWF in the pathogenesis of von Willebrand disease.

Novel splicing (c.6529-1G&gt;T) and missense (c.1667G&gt;A) mutations causing factor V deficiency

Congenital factor V deficiency (FVD) is a rare bleeding disorder. In this study, we investigated the genetic basis in an African American patient with factor V activity 3%. Custom sequence capture and targeted next-generation (NGS) sequencing of the F5 gene were undertaken followed by PCR and Sanger sequencing. Two novel variants were identified. In silico analyses correlated clinically with the patient's factor V activity and hemorrhagic tendency. A review of the literature regarding these genomic alterations is presented. We described two novel mutations causing moderate FVD. The first, Chr1:g.169483698C>A with cDNA change (F5):c.6529-1G>T, occurred in a conserved nucleotide at the canonical acceptor splice site of intron 24. The second, Chr1:g.169515775C>T with cDNA change (F5):c.1667G>A, was a missense variant of exon 11, affecting a highly conserved amino acid in the A2 domain. Further research into the mechanisms of F5 mutations leading to FVD and residual factor V expression are needed.

Cryo-EM structures of human coagulation factors V and Va

Coagulation factor V (fV) is the precursor of fVa, which, together with fXa, Ca2+, and phospholipids, defines the prothrombinase complex and activates prothrombin in the penultimate step of the coagulation cascade. We solved the cryogenic electron microscopy (cryo-EM) structures of human fV and fVa at atomic (3.3 Å) and near-atomic (4.4 Å) resolution, respectively. The structure of fV reveals the entire A1-A2-B-A3-C1-C2 assembly, but with a surprisingly disordered B domain. The C1 and C2 domains provide a platform for interaction with phospholipid membranes and support the A1 and A3 domains, with the A2 domain sitting on top of them. The B domain is highly dynamic and visible only for short segments connecting to the A2 and A3 domains. The A2 domain reveals all sites of proteolytic processing by thrombin and activated protein C, a partially buried epitope for binding fXa, and fully exposed epitopes for binding activated protein C and prothrombin. Removal of the B domain and activation to fVa exposes the sites of cleavage by activated protein C at R306 and R506 and produces increased disorder in the A1-A2-A3-C1-C2 assembly, especially in the C-terminal acidic portion of the A2 domain that is responsible for prothrombin binding. Ordering of this region and full exposure of the fXa epitope emerge as necessary steps in the assembly of the prothrombin-prothrombinase complex. These structures offer molecular context for the function of fV and fVa and pioneer the analysis of coagulation factors by cryo-EM.

Oxidation shuts down an auto-inhibitory mechanism of von Willebrand factor.

The blood protein von Willebrand factor (VWF) is a key link between inflammation and pathological thrombus formation. In particular, oxidation of methionine residues in specific domains of VWF due to the release of oxidants in inflammatory conditions has been linked to an increased platelet-binding activity. However, the atomistic details of how methionine oxidation activates VWF have not been elucidated to date. Yet understanding the activation mechanism of VWF under oxidizing conditions can lead to the development of novel therapeutics that target VWF selectively under inflammatory conditions in order to reduce its thrombotic activity while maintaining its haemostatic function. In this manuscript, we used a combination of a dynamic flow assay and molecular dynamics (MD) simulations to investigate how methionine oxidation removes an auto-inhibitory mechanism of VWF. Results from the dynamic flow assay revealed that oxidation does not directly activate the A1 domain, which is the domain in VWF that contains the binding site to the platelet surface receptor glycoprotein Ibα (GpIbα), but rather removes the inhibitory function of the neighboring A2 and A3 domains. Furthermore, the MD simulations combined with free energy perturbation calculations suggested that methionine oxidation may destabilize the binding interface between the A1 and A2 domains leading to unmasking of the GpIbα-binding site in the A1 domain.

The C‐terminal acidic region in the A1 domain of factor VIII facilitates thrombin‐catalyzed activation and cleavage at Arg372

BackgroundFactor VIII (FVIII) is activated by thrombin‐catalyzed cleavage at three sites. Previous reports indicated that the A2 domain contained thrombin‐interactive sites responsible for cleavage at Arg372. We have also found that the A1 domain of FVIII bound to the anion‐binding exosite I of thrombin. The present study focused, therefore, on thrombin interaction with A1 residues 337‐372 containing clustered acidic and hirugen‐like sequences. AimTo identify specific thrombin‐interactive site(s) within the A1 acidic region of FVIII. Methods and ResultsThe synthetic peptide of residues 337‐353 with sulfated Tyr346 (337‐353S) significantly blocked thrombin‐catalyzed FVIII activation and cleavage at Arg372, while a corresponding peptide of residues 354‐372 had no significant effect. Treatment with 1‐ethyl‐3‐(3‐dimethylaminopropyl)‐carbodiimide to cross‐link thrombin and 340‐350S suggested that the 344‐349 clustered acidic region was involved in thrombin interaction. Alanine‐substituted FVIII mutants, Y346A and D347A/D348A/D349A, depressed thrombin‐catalyzed activation and cleavage at Arg372, with peak activation at ~ 50% and cleavage rates of ~ 10% to 20% compared to wild type (WT). The peak level of thrombin‐catalyzed activation and the cleavage rate at Arg372 using FVIII mutants with 337‐346 residues substituted with hirugen‐sequences (MKNNEEAEDY337‐346GDFEEIPEEY) were ~ 1.5‐ and ~ 2.5‐fold of WT, respectively. Surface plasmon resonance‐based analysis demonstrated that the Kd for active‐site modified thrombin interactions using Y346A and D347A/D348A/D349A mutants was ~ 3‐ to 6‐fold higher than that of WT, and that the hirugen‐hybrid mutant facilitated association kinetics ~ 1.8‐fold of WT. ConclusionResidues 346‐349 with sulfated Tyr provided a thrombin‐interactive site responsible for activation and cleavage at Arg372. A hirugen‐hybrid A1 mutant showed more efficient thrombin‐catalyzed cleavage at Arg372.

Caplacizumab for Acute Thrombotic Thrombocytopenic Purpura.

Acute thrombotic thrombocytopenic purpura (aTTP) is a rare microangiopathic hemolytic anemia. Standard of care currently includes plasma exchange and immunosuppressive agents, including glucocorticoids, vincristine, and rituximab. Even with these therapies, relapse occurs in 36% of patients, and mortality ranges from 10% to 20%. Caplacizumab is a novel agent approved for the treatment of adult patients with aTTP in conjunction with plasma exchange and immunosuppressive therapies. It works by binding to the A1 domain of von Willebrand factor (VWF), blocking platelets from binding to VWF and aggregating. In clinical trials, patients who received caplacizumab compared with placebo were more likely to have a normalization of their platelet count, a lower rate of recurrence, and a lower incidence of the composite of aTTP-related death, recurrence, or major thromboembolic event. The side effect profile is rather benign and includes epistaxis, headache, and gingival bleeding. Caplacizumab is only available through specialty pharmacy services due to its high cost. Providers should be aware of and prepared for the prior authorization process required to assist their patients in gaining access to the medication. Currently, there is no formal consensus regarding caplacizumab's place in therapy for patients with aTTP, but it remains an option for refractory cases.

Thermodynamic stability of hnRNP A1 low complexity domain revealed by high-pressure NMR.

We have investigated the pressure- and temperature-induced conformational changes associated with the low complexity domain of hnRNP A1, an RNA-binding protein able to phase separate in response to cellular stress. Solution NMR spectra of the hnRNP A1 low-complexity domain fused with protein-G B1 domain were collected from 1 to 2500 bar and from 268 to 290 K. While the GB1 domain shows the typical pressure-induced and cold temperature-induced unfolding expected for small globular domains, the low-complexity domain of hnRNP A1 exhibits unusual pressure and temperature dependences. We observed that the low-complexity domain is pressure sensitive, undergoing a major conformational transition within the prescribed pressure range. Remarkably, this transition has the inverse temperature dependence of a typical folding-unfolding transition. Our results suggest the presence of a low-lying extended and fully solvated state(s) of the low-complexity domain that may play a role in phase separation. This study highlights the exquisite sensitivity of solution NMR spectroscopy to observe subtle conformational changes and illustrates how pressure perturbation can be used to determine the properties of metastable conformational ensembles.

The VWF/LRP4/αVβ3-axis represents a novel pathway regulating proliferation of human vascular smooth muscle cells.

Von Willebrand factor (VWF) is a plasma glycoprotein involved in primary haemostasis, while also having additional roles beyond haemostasis namely in cancer, inflammation, angiogenesis, and potentially in vascular smooth muscle cell (VSMC) proliferation. Here, we addressed how VWF modulates VSMC proliferation and investigated the underlying molecular pathways and the in vivo pathophysiological relevance. VWF induced proliferation of human aortic VSMCs and also promoted VSMC migration. Treatment of cells with a siRNA against αv integrin or the RGT-peptide blocking αvβ3 signalling abolished proliferation. However, VWF did not bind to αvβ3 on VSMCs through its RGD-motif. Rather, we identified the VWF A2 domain as the region mediating binding to the cells. We hypothesized the involvement of a member of the LDL-related receptor protein (LRP) family due to their known ability to act as co-receptors. Using the universal LRP-inhibitor receptor-associated protein, we confirmed LRP-mediated VSMC proliferation. siRNA experiments and confocal fluorescence microscopy identified LRP4 as the VWF-counterreceptor on VSMCs. Also co-localization between αvβ3 and LRP4 was observed via proximity ligation analysis and immuno-precipitation experiments. The pathophysiological relevance of our data was supported by VWF-deficient mice having significantly reduced hyperplasia in carotid artery ligation and artery femoral denudation models. In wild-type mice, infiltration of VWF in intimal regions enriched in proliferating VSMCs was found. Interestingly, also analysis of human atherosclerotic lesions showed abundant VWF accumulation in VSMC-proliferating rich intimal areas. VWF mediates VSMC proliferation through a mechanism involving A2 domain binding to the LRP4 receptor and integrin αvβ3 signalling. Our findings provide new insights into the mechanisms that drive physiological repair and pathological hyperplasia of the arterial vessel wall. In addition, the VWF/LRP4-axis may represent a novel therapeutic target to modulate VSMC proliferation.

Structure of blood coagulation factor VIII in complex with an anti–C1 domain pathogenic antibody inhibitor

AbstractAntibody inhibitor development in hemophilia A represents the most significant complication resulting from factor VIII (fVIII) replacement therapy. Recent studies have demonstrated that epitopes present in the C1 domain contribute to a pathogenic inhibitor response. In this study, we report the structure of a group A anti–C1 domain inhibitor, termed 2A9, in complex with a B domain–deleted, bioengineered fVIII construct (ET3i). The 2A9 epitope forms direct contacts to the C1 domain at 3 different surface loops consisting of Lys2065-Trp2070, Arg2150-Tyr2156, and Lys2110-Trp2112. Additional contacts are observed between 2A9 and the A3 domain, including the Phe1743-Tyr1748 loop and the N-linked glycosylation at Asn1810. Most of the C1 domain loops in the 2A9 epitope also represent a putative interface between fVIII and von Willebrand factor. Lastly, the C2 domain in the ET3i:2A9 complex adopts a large, novel conformational change, translocating outward from the structure of fVIII by 20 Å. This study reports the first structure of an anti–C1 domain antibody inhibitor and the first fVIII:inhibitor complex with a therapeutically active fVIII construct. Further structural understanding of fVIII immunogenicity may result in the development of more effective and safe fVIII replacement therapies.

Conformation-dependent blockage of activated VWF improves outcomes of traumatic brain injury in mice

AbstractTraumatic brain injury-induced coagulopathy (TBI-IC) causes life-threatening secondary intracranial bleeding. Its pathogenesis differs mechanistically from that of coagulopathy arising from extracranial injuries and hemorrhagic shock, but it remains poorly understood. We report results of a study designed to test the hypothesis that von Willebrand factor (VWF) released during acute TBI is intrinsically hyperadhesive because its platelet-binding A1-domain is exposed and contributes to TBI-induced vascular leakage and consumptive coagulopathy. This hyperadhesive VWF can be selectively blocked by a VWF A2-domain protein to prevent TBI-IC and to improve neurological function with a minimal risk of bleeding. We demonstrated that A2 given through intraperitoneal injection or IV infusion reduced TBI-induced death by >50% and significantly improved the neurological function of C57BL/6J male mice subjected to severe lateral fluid percussion injury. A2 protected the endothelium from extracellular vesicle-induced injury, reducing TBI-induced platelet activation and microvesiculation, and preventing a TBI-induced hypercoagulable state. A2 achieved this therapeutic efficacy by specifically blocking the A1 domain exposed on the hyperadhesive VWF released during acute TBI. These results suggest that VWF plays a causal role in the development of TBI-IC and is a therapeutic target for this life-threatening complication of TBI.

Multimodal Imaging of Synaptic Vesicles with a Single Probe

A complete understanding of synaptic vesicle recycling requires the use of multiple microscopy methods to obtain complementary information. However, many currently available probes are limited to a specific microscopy modality, which necessitates the use of multiple probes and labeling paradigms. Given the complexity of vesicle populations and recycling pathways, having new single vesicle probes that could be used for multiple microscopy techniques would complement existing sets of tools for studying vesicle function. Here we present a novel probe based on the membrane-binding C2 domain of cytosolic phospholipase A2 (cPLA2) that fulfills this need. By conjugating the C2 domain with different detectable tags, we demonstrate that a single, modular probe can allow synaptic vesicles to be imaged at multiple levels of spatial and temporal resolution.

Prioritization and characterization of validated biofilm blockers targeting glucosyltransferase C of Streptococcus mutans

To date, several Glucosyltransferase C (GtfC) inhibitors have been identified and experimentally validated. All these inhibitors have been validated at different experimental conditions like degree of purity, animal models, kinetic conditions, experimental environment etc.; and most of these inhibitors (ligands) proved to be quite effective in their respective validation environment. However, due to varied experimental validation conditions, and absence of molecular interaction data, there is no way to prioritize these validated ligands for their inhibition potential against GtfC. The present study is a novel attempt of comparative evaluation of the interaction of the validated ligands on a single platform and under similar conditions with a dual objective, i.e. ligand prioritization for their respective inhibitory potential and elucidation of the involved unknown molecular interactions. Carbohydrate derivatives (6-Deoxy sucrose and Trichloro-galactosucrose) were identified as the most promising GtfC inhibitors. In addition, Asp588, Trp517, and Asn481 amino acid residues of the domain A1 proved vital for the inhibitory effect. The study highlights the importance of the comparative analysis of the validated ligands in order to identify the most promising leads for drug discovery against dental caries.

The Use of Fluorescently Labeled ARC1779 Aptamer for Assessing the Effect of H2O2 on von Willebrand Factor Exocytosis

Here, we propose a new approach for quantitative estimation of von Willebrand factor (vWF) exposed on the surface of endothelial cells (ECs) using the ARC1779 aptamer that interacts with the vWF A1 domain. To visualize complex formation between vWF and the aptamer, the latter was conjugated with the Cy5 fluorescent label. Cultured human umbilical vein endothelial cells (HUVEC) were stained with the ARC1779-Cy5 conjugate and imaged with a fluorescence microscope. The images were analyzed with the CellProfiler software. vWF released from the Weibel–Palade bodies was observed as bright dot-like structures of round and irregular shape, the number of which increased several times after HUVEC exposure to histamine or thrombin. Staining with ARC1779-Cy5 also revealed long filamentous vWF structures on the surface of activated HUVEC. vWF secretion by ECs is activated by the second messengers cAMP and Ca2+. There is evidence that hydrogen peroxide also acts as a second messenger in ECs. In addition, exogenous H2O2 formed in leukocytes can enter ECs. The aim of our study was to determine the effect of H2O2 on the vWF exposure at the surface of HUVEC using the proposed method. It was shown that hydrogen peroxide at concentration 100 µM, which is lower than the cytotoxicity threshold of H2O2 for cultured HUVEC, increased several times the number of dot-like structures and total amount of vWF exposed on plasma membrane of HUVEC, which suggest that H2O2 acts as a mediator that activates exocytosis of Weibel–Palade bodies and vWF secretion in the vascular endothelium during inflammation and upon elevated generation of endogenous reactive oxygen species in ECs.

The VP1u of Human Parvovirus B19: A Multifunctional Capsid Protein with Biotechnological Applications.

The viral protein 1 unique region (VP1u) of human parvovirus B19 (B19V) is a multifunctional capsid protein with essential roles in virus tropism, uptake, and subcellular trafficking. These functions reside on hidden protein domains, which become accessible upon interaction with cell membrane receptors. A receptor-binding domain (RBD) in VP1u is responsible for the specific targeting and uptake of the virus exclusively into cells of the erythroid lineage in the bone marrow. A phospholipase A2 domain promotes the endosomal escape of the incoming virus. The VP1u is also the immunodominant region of the capsid as it is the target of neutralizing antibodies. For all these reasons, the VP1u has raised great interest in antiviral research and vaccinology. Besides the essential functions in B19V infection, the remarkable erythroid specificity of the VP1u makes it a unique erythroid cell surface biomarker. Moreover, the demonstrated capacity of the VP1u to deliver diverse cargo specifically to cells around the proerythroblast differentiation stage, including erythroleukemic cells, offers novel therapeutic opportunities for erythroid-specific drug delivery. In this review, we focus on the multifunctional role of the VP1u in B19V infection and explore its potential in diagnostics and erythroid-specific therapeutics.

Hydrogen-Deuterium Exchange Mass Spectrometry Identifies Activated Factor IX-Induced molecular Changes in Activated Factor VIII.

Hydrogen-deuterium exchange mass spectrometry (HDX-MS) was employed to gain insight into the changes in factor VIII (FVIII) that occur upon its activation and assembly with activated factor IX (FIXa) on phospholipid membranes. HDX-MS analysis of thrombin-activated FVIII (FVIIIa) revealed a marked increase in deuterium incorporation of amino acid residues along the A1-A2 and A2-A3 interface. Rapid dissociation of the A2 domain from FVIIIa can explain this observation. In the presence of FIXa, enhanced deuterium incorporation at the interface of FVIIIa was similar to that of FVIII. This is compatible with the previous finding that FIXa contributes to A2 domain retention in FVIIIa. A2 domain region Leu631-Tyr637, which is not part of the interface between the A domains, also showed a marked increase in deuterium incorporation in FVIIIa compared with FVIII. Deuterium uptake of this region was decreased in the presence of FIXa beyond that observed in FVIII. This implies that FIXa alters the conformation or directly interacts with this region in FVIIIa. Replacement of Val634 in FVIII by alanine using site-directed mutagenesis almost completely impaired the ability of the activated cofactor to enhance the activity of FIXa. Surface plasmon resonance analysis revealed that the rates of A2 domain dissociation from FVIIIa and FVIIIa-Val634Ala were indistinguishable. HDX-MS analysis showed, however, that FIXa was unable to retain the A2 domain in FVIIIa-Val634Ala. The combined results of this study suggest that the local structure of Leu631-Tyr637 is altered by FIXa and that this region contributes to the cofactor function of FVIII.

CGTase, a novel antimicrobial protein from Bacillus cereus YUPP-10, suppresses Verticillium dahliae and mediates plant defence responses.

Verticillium wilt is a plant vascular disease caused by the soilborne fungus Verticillium dahliae that severely limits cotton production. In a previous study, we screened Bacillus cereus YUPP‐10, an efficient antagonistic bacterium, to uncover mechanisms for controlling verticillium wilt. Here, we report a novel antimicrobial cyclodextrin glycosyltransferase (CGTase) from YUPP‐10. Compared to other CGTases, six different conserved domains were identified, and six mutants were constructed by gene splicing with overlap extension PCR. Functional analysis showed that domain D was important for hydrolysis activity and domains A1 and C were important for inducing disease resistance. Direct effects of recombinant CGTase on V. dahliae included reduced mycelial growth, spore germination, spore production, and microsclerotia germination. In addition, CGTase also elicited cotton's innate defence reactions. Transgenic Arabidopsis thaliana lines that overexpress CGTase showed higher resistance to verticillium wilt. Transgenic CGTase A. thaliana plants grew faster and resisted disease better. CGTase overexpression enabled a burst of reactive oxygen species production and activated pathogenesis‐related gene expression, indicating that the transgenic cotton was better prepared to protect itself from infection. Our work revealed that CGTase could inhibit the growth of V. dahliae, activate innate immunity, and play a major role in the biocontrol of fungal pathogens.