The 3CL protease, a highly conserved enzyme in the coronavirus, plays a crucial role in the viral life cycle by facilitating viral replication through precise cleavage of polyproteins. Beyond its proteolytic function,...The 3CL protease, a highly conserved enzyme in the coronavirus, plays a crucial role in the viral life cycle by facilitating viral replication through precise cleavage of polyproteins. Beyond its proteolytic function, the 3CL protease also engages in intricate interactions with host cell proteins involved in critical cellular processes such as transcription, translation, and nuclear-cytoplasmic transport, effectively hijacking cellular machinery to promote viral replication. Additionally, it disrupts innate immune signaling pathways, suppresses interferon activity and cleaves antiviral proteins. Furthermore, it modulates host cell death pathways including pyroptosis and apoptosis, interferes with autophagy and inhibits stress granule formation to maintain viral infection and exacerbate viral pathogenesis. This review highlights the molecular mechanisms by which the 3CL protease orchestrates virus-host interactions, emphasizing its central role in coronavirus pathogenesis and highlighting potential therapeutic targets for future interventions.展开更多
Porcine epidemic diarrhea (PED), caused by porcine epidemic diarrhea virus (PEDV), is a highly contagious gastrointestinal disease characterized by vomiting, diarrhea, and dehydration, with mortality rates approaching...Porcine epidemic diarrhea (PED), caused by porcine epidemic diarrhea virus (PEDV), is a highly contagious gastrointestinal disease characterized by vomiting, diarrhea, and dehydration, with mortality rates approaching 100% among suckling piglets. The PEDV 3C-like protease (3CLpro) is essential for viral replication and regarded as a critical target for antiviral inhibitor development. In this study, we aimed to identify small-molecule inhibitors of PEDV by targeting 3CLpro. Virtual screening of 1.6 million compounds from the ChemDiv library identified four potential candidates. Molecular dynamics simulations, specifically analyzing RMSD, RMSF, and Rg, demonstrated increased structural stability of the compound-protease complexes compared to the monomeric enzyme. All compounds had low cytotoxicity in Vero cells (CC_(50) > 200 μM). Fluorescence resonance energy transfer-based assays demonstrated dose-dependent inhibitory activity of the compounds against 3CLpro. Among the candidates, compound F366-0161 exhibited the weakest inhibition, with an IC_(50) value of 151.5 μM. Two analogues, 3238-0395 (IC_(50) of 121.4 μM) and L878-0493 (IC_(50) of 123.6 μM), exhibited moderately enhanced activity. Y041-1672 was identified as the most effective inhibitor, with an IC_(50) of 86.48 μM. In viral replication inhibition assays, Y041-1672 reduced PEDV replication, with an EC_(50) of 17.97 μM and a selectivity index (SI) of 15.5 (CC_(50) /EC_(50) ). These results were validated by RT-qPCR, plaque assays, immunofluorescence, and Western blot analyses. In vitro validation confirmed Y041-1672 as the optimal antiviral candidate, and time-of-addition experiments indicated that inhibition primarily occurred during viral replication. This study identifies scaffold molecules for PEDV antiviral drug development, providing strategic insights for PED treatment.展开更多
The CX3CL1/CX3CR1 signaling axis is established as a pivotal regulator in the pathogenesis of atherosclerosis,with well-documented roles in orchestrating inflammatory responses,mediating immune cell recruitment,and in...The CX3CL1/CX3CR1 signaling axis is established as a pivotal regulator in the pathogenesis of atherosclerosis,with well-documented roles in orchestrating inflammatory responses,mediating immune cell recruitment,and influencing vascular remodeling.This review provides a comprehensive synthesis of current knowledge regarding the structural characteristics and functional properties of the CX3CL1/CX3CR1 pathway.This study delves into its specific mechanistic contributions to atherosclerosis,placing particular emphasis on its regulatory influence across diverse cell types,including arterial endothelial cells,smooth muscle cells,and macrophages.Furthermore,the pathway’s integral involvement in both the initiation and progression of atherosclerotic plaques is dissected,highlighting its critical impact on plaque stability and susceptibility to rupture.The review also extends to the pathogenic significance of CX3CL1/CX3CR1 signaling in atherosclerosis-related comorbidities,incorporating the latest advancements in understanding its roles in coronary heart disease,stroke,and other cardiovascular disorders.By critically integrating findings from the extant literature,this review constructs a foundational framework to guide future investigations and underscores the substantial translational potential of targeting this pathway for therapeutic intervention in clinical settings.展开更多
Severe acute respiratory syndrome coronavirus 2(SARS-CoV-2),the causative agent of novel coronavirus disease 2019,can cause acute respiratory symptoms and even death globally.However,the immune escape mechanism and vi...Severe acute respiratory syndrome coronavirus 2(SARS-CoV-2),the causative agent of novel coronavirus disease 2019,can cause acute respiratory symptoms and even death globally.However,the immune escape mechanism and viral pathogenesis remain poorly understood.Here,we report that the SARS-CoV-23C-like(3CL)protease specifically cleaves gasdermin D(GSDMD)at Q29 and Q193,producing two N-terminal fragments,GSDMD1-29 and GSDMD1-193.We also found that SARS-CoV-2 infection induced the cleavage of GSDMD.Then,we demonstrated that the ability to cleave GSDMD was dependent on the protease activity of the 3CL protease.Interestingly,unlike the GSDMD1-275 fragment cleaved by caspase-1,GSDMD1-29 and GSDMD1-193 did not trigger pyroptosis or inhibit SARS-CoV-2 replication.Additionally,various RNA viral proteases display different preferences for cleaving GSDMD at Q29 and Q193.Our findings reveal a mechanism by which SARS-CoV-2 and other RNA viruses inhibit pyroptosis,highlighting the critical role of the 3CL protease in immune evasion and viral replication.展开更多
Ferroptosis has been shown to mediate the development of fibrosis.Polyphyllin VII(PP7),a bioactive component of Paris polyphylla,exhibits potent anti-inflammatory activity and can significantly alleviate liver fibrosi...Ferroptosis has been shown to mediate the development of fibrosis.Polyphyllin VII(PP7),a bioactive component of Paris polyphylla,exhibits potent anti-inflammatory activity and can significantly alleviate liver fibrosis.In this study,treatment with PP7 significantly inhibited the proliferation and activation of hepatic stellate cells(HSCs),which could be suppressed by a ferroptosis inhibitor.In addition,it promoted HSC ferroptosis by suppressing glutathione(GSH)peroxidase 4(GPX4)and enhanced the expression of CX3C chemokine ligand 1(CX3CL1).Depletion of CX3CL1 attenuated the effects of PP7 on the activation and ferroptosis of HSCs and the expression of GPX4.Notably,CX3CL1 directly interacted with GPX4,triggering HSC ferroptosis.The transcription factor hypermethylated in cancer 1(Hic1),which binds to the Cx3cl1 promoter,increased the expression of CX3CL1.Its absence resulted in downregulation of CX3CL1,suppressing the GPX4-dependent ferroptosis of PP7-treated HSCs and promoting their activation.HIC1 was found to directly interact with PP7 at the GLY164 site.Co-culture experiments showed that PP7-induced HSC ferroptosis attenuated macrophage recruitment by regulating inflammation-related genes.HSC-specific inhibition of HIC1 counteracted PP7-induced collagen depletion and HSC ferroptosis in vivo.These findings suggest that PP7 induces HSC ferroptosis through the HIC1/CX3CL1/GPX4 axis.展开更多
为了探讨复合物中分子间相互作用对单体分子动力学的影响,对CF3Cl-CO复合物中单体振动谱带基线平移进行了预测。首先,采用二阶多体微扰理论(M?ller-Plesset perturbation theory of second order, MP2)结合扩展相关一致性极化价三重ζ基...为了探讨复合物中分子间相互作用对单体分子动力学的影响,对CF3Cl-CO复合物中单体振动谱带基线平移进行了预测。首先,采用二阶多体微扰理论(M?ller-Plesset perturbation theory of second order, MP2)结合扩展相关一致性极化价三重ζ基组(augmented correlation-consistent polarized valence triple-zeta, Aug-cc-pVTZ)对该复合物的5种结构(S1′、S2、S3、S4、A)进行几何优化、频率计算;然后,引入键函数(3s3p2d1f),在包含单激发、双激发和微扰三激发的耦合簇(coupled cluster with single, double, and perturbative triple excitations, CCSD(T))方法结合Aug-cc-pVTZ的水平上计算稳定结构的分子间相互作用能,并对最稳定结构的单体谱带基线平移进行预测。结果表明,S1′为该复合物的最稳定结构,其分子间相互作用能为-3.81 kJ/mol,其能量分解中交换能占比最高为38%。此外,研究中发现了复合物的2个新的稳定结构S4、A,其分子间相互作用能分别为-2.68、-2.72 kJ/mol。单体谱带基线平移预测表明,在S1′结构中,相比于自由单体振动,复合物中CF3Cl单体的C-Cl键伸缩振动发生了红移,而CO的伸缩振动则表现为蓝移,这表明分子间卤键对分子内动力学存在显著影响。该研究结果为进一步实验测量CF3Cl-CO复合物光谱提供了理论参考。展开更多
基金supported by the National Natural Science Foundation of China(grant no.82370015).
文摘The 3CL protease, a highly conserved enzyme in the coronavirus, plays a crucial role in the viral life cycle by facilitating viral replication through precise cleavage of polyproteins. Beyond its proteolytic function, the 3CL protease also engages in intricate interactions with host cell proteins involved in critical cellular processes such as transcription, translation, and nuclear-cytoplasmic transport, effectively hijacking cellular machinery to promote viral replication. Additionally, it disrupts innate immune signaling pathways, suppresses interferon activity and cleaves antiviral proteins. Furthermore, it modulates host cell death pathways including pyroptosis and apoptosis, interferes with autophagy and inhibits stress granule formation to maintain viral infection and exacerbate viral pathogenesis. This review highlights the molecular mechanisms by which the 3CL protease orchestrates virus-host interactions, emphasizing its central role in coronavirus pathogenesis and highlighting potential therapeutic targets for future interventions.
基金supported by the National Key Research and Development Program of China(2021YFD1800303)the National Natural Science Foundation of China(32473044).
文摘Porcine epidemic diarrhea (PED), caused by porcine epidemic diarrhea virus (PEDV), is a highly contagious gastrointestinal disease characterized by vomiting, diarrhea, and dehydration, with mortality rates approaching 100% among suckling piglets. The PEDV 3C-like protease (3CLpro) is essential for viral replication and regarded as a critical target for antiviral inhibitor development. In this study, we aimed to identify small-molecule inhibitors of PEDV by targeting 3CLpro. Virtual screening of 1.6 million compounds from the ChemDiv library identified four potential candidates. Molecular dynamics simulations, specifically analyzing RMSD, RMSF, and Rg, demonstrated increased structural stability of the compound-protease complexes compared to the monomeric enzyme. All compounds had low cytotoxicity in Vero cells (CC_(50) > 200 μM). Fluorescence resonance energy transfer-based assays demonstrated dose-dependent inhibitory activity of the compounds against 3CLpro. Among the candidates, compound F366-0161 exhibited the weakest inhibition, with an IC_(50) value of 151.5 μM. Two analogues, 3238-0395 (IC_(50) of 121.4 μM) and L878-0493 (IC_(50) of 123.6 μM), exhibited moderately enhanced activity. Y041-1672 was identified as the most effective inhibitor, with an IC_(50) of 86.48 μM. In viral replication inhibition assays, Y041-1672 reduced PEDV replication, with an EC_(50) of 17.97 μM and a selectivity index (SI) of 15.5 (CC_(50) /EC_(50) ). These results were validated by RT-qPCR, plaque assays, immunofluorescence, and Western blot analyses. In vitro validation confirmed Y041-1672 as the optimal antiviral candidate, and time-of-addition experiments indicated that inhibition primarily occurred during viral replication. This study identifies scaffold molecules for PEDV antiviral drug development, providing strategic insights for PED treatment.
基金Study on the Protective Mechanism of Safflower Yellow Pigment on Vascular Endothelial Function in Patients with Phlegm-Stasis Syndrome Coronary Heart Disease and Stable Angina Pectoris(Project No.20222A010012)Single-Cell Immune Panorama Study on Neiguan(PC6)Acupoint Injection for Improving Pyroptosis in Chronic Heart Failure(Project No.2025A03J3499)+1 种基金The study on the effect and mechanism of Guanxinning Tablets on vascular homeostasis and remodeling in populations with early vascular aging(Project No.2024QC-B1007)Guangzhou Key Laboratory of Traditional Chinese Medicine Rehabilitation(Project No.2024A03J0790).
文摘The CX3CL1/CX3CR1 signaling axis is established as a pivotal regulator in the pathogenesis of atherosclerosis,with well-documented roles in orchestrating inflammatory responses,mediating immune cell recruitment,and influencing vascular remodeling.This review provides a comprehensive synthesis of current knowledge regarding the structural characteristics and functional properties of the CX3CL1/CX3CR1 pathway.This study delves into its specific mechanistic contributions to atherosclerosis,placing particular emphasis on its regulatory influence across diverse cell types,including arterial endothelial cells,smooth muscle cells,and macrophages.Furthermore,the pathway’s integral involvement in both the initiation and progression of atherosclerotic plaques is dissected,highlighting its critical impact on plaque stability and susceptibility to rupture.The review also extends to the pathogenic significance of CX3CL1/CX3CR1 signaling in atherosclerosis-related comorbidities,incorporating the latest advancements in understanding its roles in coronary heart disease,stroke,and other cardiovascular disorders.By critically integrating findings from the extant literature,this review constructs a foundational framework to guide future investigations and underscores the substantial translational potential of targeting this pathway for therapeutic intervention in clinical settings.
基金supported by the National Natural Science Foundation of China(82370015).
文摘Severe acute respiratory syndrome coronavirus 2(SARS-CoV-2),the causative agent of novel coronavirus disease 2019,can cause acute respiratory symptoms and even death globally.However,the immune escape mechanism and viral pathogenesis remain poorly understood.Here,we report that the SARS-CoV-23C-like(3CL)protease specifically cleaves gasdermin D(GSDMD)at Q29 and Q193,producing two N-terminal fragments,GSDMD1-29 and GSDMD1-193.We also found that SARS-CoV-2 infection induced the cleavage of GSDMD.Then,we demonstrated that the ability to cleave GSDMD was dependent on the protease activity of the 3CL protease.Interestingly,unlike the GSDMD1-275 fragment cleaved by caspase-1,GSDMD1-29 and GSDMD1-193 did not trigger pyroptosis or inhibit SARS-CoV-2 replication.Additionally,various RNA viral proteases display different preferences for cleaving GSDMD at Q29 and Q193.Our findings reveal a mechanism by which SARS-CoV-2 and other RNA viruses inhibit pyroptosis,highlighting the critical role of the 3CL protease in immune evasion and viral replication.
基金supported by the National Natural Science Foundation of China(Grant No.:81873576)Wenzhou Municipal Science and Technology Bureau,China(Grant No.:Y20220023).
文摘Ferroptosis has been shown to mediate the development of fibrosis.Polyphyllin VII(PP7),a bioactive component of Paris polyphylla,exhibits potent anti-inflammatory activity and can significantly alleviate liver fibrosis.In this study,treatment with PP7 significantly inhibited the proliferation and activation of hepatic stellate cells(HSCs),which could be suppressed by a ferroptosis inhibitor.In addition,it promoted HSC ferroptosis by suppressing glutathione(GSH)peroxidase 4(GPX4)and enhanced the expression of CX3C chemokine ligand 1(CX3CL1).Depletion of CX3CL1 attenuated the effects of PP7 on the activation and ferroptosis of HSCs and the expression of GPX4.Notably,CX3CL1 directly interacted with GPX4,triggering HSC ferroptosis.The transcription factor hypermethylated in cancer 1(Hic1),which binds to the Cx3cl1 promoter,increased the expression of CX3CL1.Its absence resulted in downregulation of CX3CL1,suppressing the GPX4-dependent ferroptosis of PP7-treated HSCs and promoting their activation.HIC1 was found to directly interact with PP7 at the GLY164 site.Co-culture experiments showed that PP7-induced HSC ferroptosis attenuated macrophage recruitment by regulating inflammation-related genes.HSC-specific inhibition of HIC1 counteracted PP7-induced collagen depletion and HSC ferroptosis in vivo.These findings suggest that PP7 induces HSC ferroptosis through the HIC1/CX3CL1/GPX4 axis.
文摘为了探讨复合物中分子间相互作用对单体分子动力学的影响,对CF3Cl-CO复合物中单体振动谱带基线平移进行了预测。首先,采用二阶多体微扰理论(M?ller-Plesset perturbation theory of second order, MP2)结合扩展相关一致性极化价三重ζ基组(augmented correlation-consistent polarized valence triple-zeta, Aug-cc-pVTZ)对该复合物的5种结构(S1′、S2、S3、S4、A)进行几何优化、频率计算;然后,引入键函数(3s3p2d1f),在包含单激发、双激发和微扰三激发的耦合簇(coupled cluster with single, double, and perturbative triple excitations, CCSD(T))方法结合Aug-cc-pVTZ的水平上计算稳定结构的分子间相互作用能,并对最稳定结构的单体谱带基线平移进行预测。结果表明,S1′为该复合物的最稳定结构,其分子间相互作用能为-3.81 kJ/mol,其能量分解中交换能占比最高为38%。此外,研究中发现了复合物的2个新的稳定结构S4、A,其分子间相互作用能分别为-2.68、-2.72 kJ/mol。单体谱带基线平移预测表明,在S1′结构中,相比于自由单体振动,复合物中CF3Cl单体的C-Cl键伸缩振动发生了红移,而CO的伸缩振动则表现为蓝移,这表明分子间卤键对分子内动力学存在显著影响。该研究结果为进一步实验测量CF3Cl-CO复合物光谱提供了理论参考。
