查看更多>>摘要：CRISPR-Cas7-11 is a Type Ⅲ-E CRISPR-associated nuclease that functions as a potent RNA editing tool.Tetratrico-peptide repeat fused with Cas/HEF1-associated signal transducer(TPR-CHAT)acts as a regulatory protein that interacts with CRISPR RNA(crRNA)-bound Cas7-11 to form a CRISPR-guided caspase complex(Craspase).However,the precise modulation of Cas7-11's nuclease activity by TPR-CHAT to enhance its utility requires further study.Here,we report cryo-electron microscopy(cryo-EM)structures of Desulfonema ishimotonii(Di)Cas7-11-crRNA,complexed with or without the full length or the N-terminus of TPR-CHAT.These structures unveil the molecular features of the Craspase complex.Structural analysis,combined with in vitro nuclease assay and electrophoretic mobility shift assay,reveals that DiTPR-CHAT negatively regulates the activity of DiCas7-11 by preventing target RNA from binding through the N-terminal 65 amino acids of DiTPR-CHAT(DiTPR-CHATNTD).Our work demonstrates that DiTPR-CHATNTD can function as a small unit of DiCas7-11 regulator,potentially enabling safe applications to prevent overcutting and off-target effects of the CRISPR-Cas7-11 system.
查看更多>>摘要：Almost all the neutralizing antibodies targeting the receptor-binding domain(RBD)of spike(S)protein show weakened or lost efficacy against severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)emerged or emerging variants,such as Omicron and its sub-variants.This suggests that highly conserved epitopes are crucial for the development of neutralizing antibodies.Here,we present one nanobody,N235,displaying broad neutralization against the SARS-CoV-2 prototype and multiple variants,including the newly emerged Omicron and its sub-variants.Cryo-electron microscopy demonstrates N235 binds a novel,conserved,cryptic epitope in the N-terminal domain(NTD)of the S protein,which interferes with the RBD in the neighboring S protein.The neutralization mechanism interpreted via flow cytometry and Western blot shows that N235 appears to induce the S1 subunit shedding from the trimeric S complex.Furthermore,a nano-IgM construct(MN235),engineered by fusing N235 with the human IgM Fc region,displays prevention via inducing S1 shedding and cross-linking virus particles.Compared to N235,MN235 exhibits varied enhancement in neutralization against pseudotyped and authentic viruses in vitro.The intranasal administration of MN235 in low doses can effectively prevent the infection of Omicron sub-variant BA.1 and XBB in vivo,suggesting that it can be developed as a promising prophylactic antibody to cope with the ongoing and future infection.
查看更多>>摘要：The safety and efficacy of COVID-19 vaccines in the elderly,a high-risk group for severe COVID-19 infection,have not been fully understood.To clarify these issues,this prospective study followed up 157 elderly and 73 young participants for 16 months and compared the safety,immunogenicity,and efficacy of two doses of the inactivated vaccine BBlBP-CorV followed by a booster dose of the recombinant protein vaccine ZF2001.The results showed that this vaccination protocol was safe and tolerable in the elderly.After administering two doses of the BBIBP-CorV,the positivity rates and titers of neutralizing and anti-RBD antibodies in the elderly were significantly lower than those in the young individuals.After the ZF2001 booster dose,the antibody-positive rates in the elderly were comparable to those in the young;however,the antibody titers remained lower.Gender,age,and underlying diseases were independently associated with vaccine immunogenicity in elderly individuals.The pseudovirus neutralization assay showed that,compared with those after receiving two doses of BBlBP-CorV priming,some participants obtained immunological protection against BA.5 and BF.7 after receiving the ZF2001 booster.Breakthrough infection symptoms last longer in the infected elderly and pre-infection antibody titers were negatively associated with the severity of post-infection symptoms.The antibody levels in the elderly increased significantly after breakthrough infection but were still lower than those in the young.Our data suggest that multiple booster vaccinations at short intervals to maintain high antibody levels may be an effective strategy for protecting the elderly against COVID-19.
查看更多>>摘要：Sirtuin 3(SIRT3)is well known as a conserved nicotinamide adenine dinucleotide+(NAD+)-dependent deacetylase located in the mitochondria that may regulate oxidative stress,catabolism and ATP production.Accumulating evidence has recently revealed that SIRT3 plays its critical roles in cardiac fibrosis,myocardial fibrosis and even heart failure(HF),through its deacetylation modifications.Accordingly,discovery of SIRT3 activators and elucidating their underlying mechanisms of HF should be urgently needed.Herein,we identified a new small-molecule activator of SIRT3(named 2-APQC)by the structure-based drug designing strategy.2-APQC was shown to alleviate isoproterenol(ISO)-induced cardiac hypertrophy and myocardial fibrosis in vitro and in vivo rat models.Importantly,in SIRT3 knockout mice,2-APQC could not relieve HF,suggesting that 2-APQC is dependent on SIRT3 for its protective role.Mechanically,2-APQC was found to inhibit the mammalian target of rapamycin(mTOR)-p70 ribosomal protein S6 kinase(p70S6K),c-Jun N-terminal kinase(JNK)and transforming growth factor-β(TGF-β)/small mother against decapentaplegic 3(Smad3)pathways to improve ISO-induced cardiac hypertrophy and myocardial fibrosis.Based upon RNA-seq analyses,we demonstrated that SIRT3-pyrroline-5-carboxylate reductase 1(PYCR1)axis was closely assolated with HF.By activating PYCR1,2-APQC was shown to enhance mitochondrial proline metabolism,inhibited reactive oxygen species(ROS)-p38 mitogen activated protein kinase(p38MAPK)pathway and thereby protecting against ISO-induced mitochondrialoxidative damage.Moreover,activation of SIRT3 by 2-APQC could facilitate AMP-activated protein kinase(AMPK)-Parkln axis to inhibit ISO-induced necrosis.Together,our results demonstrate that 2-APQC is a targeted SIRT3 activator that alleviates myocardial hypertrophy and fibrosis by regulating mitochondrial homeostasis,which may provide a new clue on exploiting a promising drug candidate for the future HF therapeutics.
查看更多>>摘要：Anti-PD-1 antibodies are a favorable treatment for relapsed or refractory extranodal natural killer T cell lymphoma(RR-ENKTL),however,the complete response(CR)rate and the duration of response(DOR)need to be improved.This phase 1 b/2 study investigated the safety and efficacy of sintilimab,a fully human anti-PD-1 antibody,plus chidamide,an oral subtype-selective histone deacetylase inhibitor in 38 patients with RR-ENKTL.Expected objective response rate(ORR)of combination treatment was 80％.Patients received escalating doses of chidamide,administered concomitantly with fixed-dose sintilimab in 21-days cycles up to 12 months.No dose-limiting events were observed,RP2D of chidamide was 30 mg twice a week.Twenty-nine patients were enrolled in phase 2.In the intention-to-treat population(n=37),overall response rate was 59.5％with a complete remission rate of 48.6％.The median DOR,progression-free survival(PFS),and overall survival(OS)were 25.3,23.2,and 32.9 months,respectively.The most common grade 3 or higher treatment-emergent adverse events(AEs)were neutropenia(28.9％)and thrombocytopenia(10.5％),immune-related AEs were reported in 18(47.3％)patients.Exploratory biomarker assessment suggested that a combination of dynamic plasma ctDNA and EBV-DNA played a vital prognostic role.STAT3 mutation shows an unfavorable prognosis.Although outcome of anticipate ORR was not achieved,sintilimab plus chidamide was shown to have a manageable safety profile and yielded encouraging CR rate and DOR in RR-ENKTL for the first time.It is a promising therapeutic option for this population.
查看更多>>摘要：DEAD-box helicase 17(DDX17)is a typical member of the DEAD-box family with transcriptional cofactor activity.Although DDX17 is abundantly expressed in the myocardium,its role in heart is not fully understood.We generated cardiomyocyte-specific Ddx17-knockout mice(Ddx17-cKO),cardiomyocyte-specific Ddx17 transgenic mice(Ddx17-Tg),and various models of cardiomyocyte injury and heart failure(HF).DDX17 is downregulated in the myocardium of mouse models of heart failure and cardiomyocyte injury.Cardiomyocyte-specific knockout of Ddx17 promotes autophagic flux blockage and cardiomyocyte apoptosis,leading to progressive cardiac dysfunction,maladaptive remodeling and progression to heart failure.Restoration of DDX17 expression in cardiomyocytes protects cardiac function under pathological conditions.Further studies showed that DDX17 can bind to the transcriptional repressor B-cell lymphoma 6(BCL6)and inhibit the expression of dynamin-related protein 1(DRP1).When DDX17 expression is reduced,transcriptional repression of BCL6 is attenuated,leading to increased DRP1 expression and mitochondrial fission,which in turn leads to impaired mitochondrial homeostasis and heart failure.We also investigated the correlation of DDX17 expression with cardiac function and DRP1 expression in myocardial biopsy samples from patients with heart failure.These findings suggest that DDX17 protects cardiac function by promoting mitochondrial homeostasis through the BCL6-DRP1 pathway in heart failure.
查看更多>>摘要：The immunoprotective components control COVID-19 disease severity,as well as long-term adaptive immunity maintenance and subsequent reinfection risk discrepancies across initial COVID-19 severity,remain unclarified.Here,we longitudinally analyzed SARS-CoV-2-specific immune effectors during the acute infection and convalescent phases of 165 patients with COVID-19 categorized by severity.We found that early and robust SARS-CoV-2-specific CD4+and CD8+T cell responses ameliorate disease progression and shortened hospital stay,while delayed and attenuated virus-specific CD8+T cell responses are prominent severe COVID-19 features.Delayed antiviral antibody generation rather than titer level associates with severe outcomes.Conversely,initial COVID-19 severity imprints the long-term maintenance of SARS-CoV-2-specific adaptive immunity,demonstrating that severe convalescents exhibited more sustained virus-specific antibodies and memory T cell responses compared to mild/moderate counterparts.Moreover,initial COVID-19 severity inversely correlates with SARS-CoV-2 reinfection risk.Overall,our study unravels the complicated interaction between temporal characteristics of virus-specific T cell responses and COVID-19 severity to guide future SARS-CoV-2 wave management.
查看更多>>摘要：Conventional type 1 dendritic cells(cDC1)are the essential antigen-presenting DC subset in antitumor immunity.Suppressing B-cell lymphoma 9 and B-cell lymphoma 9-like(BCL9/BCL9L)inhibits tumor growth and boosts immune responses against cancer.However,whether oncogenic BCL9/BCL9L impairs antigen presentation in tumors is still not completely understood.Here,we show that targeting BCL9/BCL9L enhanced antigen presentation by stimulating cDC1 activation and infiltration into tumor.Pharmacological inhibition of BCL9/BCL9L with a novel inhibitor hsBCL9z96 or Bcl9/Bcl9l knockout mice markedly delayed tumor growth and promoted antitumor CD8+T cell responses.Mechanistically,targeting BCL9/BCL9L promoted antigen presentation in tumors.This is due to the increase of cDC1 activation and tumor infiltration by the XCL1-XCR1 axis.Importantly,using single-cell transcriptomics analysis,we found that Bcl9/Bcl9l deficient cDC1 were superior to wild-type(WT)cDC1 at activation and antigen presentation via NF-κB/IRF1 signaling.Together,we demonstrate that targeting BCL9/BCL9L plays a crucial role in cDC1-modulated antigen presentation of tumor-derived antigens,as well as CD8+T cell activation and tumor infiltration.Targeting BCL9/BCL9L to regulate cDC1 function and directly orchestrate a positive feedback loop necessary for optimal antitumor immunity could serve as a potential strategy to counter immune suppression and enhance cancer immunotherapy.
查看更多>>摘要：Previous studies through targeted mutagenesis of K-D-K-E motif have demonstrated that 2'-O-MTase activity is essential for efficient viral replication and immune evasion.However,the K-D-K-E catalytic motif of 2'-O-MTase is highly conserved across numerous viruses,including flaviviruses,vaccinia viruses,coronaviruses,and extends even to mammals.Here,we observed a stronger 2'-O-MTase activity in SARS-CoV-2 compared to SARS-CoV,despite the presence of a consistently active catalytic center.We further identified critical residues(Leu-36,Asn-138 and lle-153)which served as determinants of discrepancy in 2'-O-MTase activity between SARS-CoV-2 and SARS-CoV.These residues significantly enhanced the RNA binding affinity of 2'-O-MTase and boosted its versatility toward RNA substrates.Of interest,a triple substitution(Leu36 → Ile36,Asn138 → His138,Ile153 → Leu153,from SARS-CoV-2 to SARS-CoV)within nsp16 resulted in a proportional reduction in viral 2'-O-methylation and impaired viral replication.Furthermore,it led to a significant upregulation of type Ⅰ interferon(IFN-Ⅰ)and proinflammatory cytokines both in vitro and vivo,relying on the cooperative sensing of melanoma differentiation-associated protein 5(MDA5)and laboratory of genetics and physiology 2(LGP2).In conclusion,our findings demonstrated that alterations in residues other than K-D-K-E of 2'-O-MTase may affect viral replication and subsequently influence pathogenesis.Monitoring changes in nsp16 residues is crucial as it may aid in identifying and assessing future alteration in viral pathogenicity resulting from natural mutations occurring in nsp16.
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