RESUMO
The coronavirus disease 2019 (COVID-19) pandemic continues worldwide with many variants arising, some of which are variants of concern (VOCs). A recent VOC, omicron (B.1.1.529), which obtains a large number of mutations in the receptor-binding domain (RBD) of the spike protein, has risen to intense scientific and public attention. Here, we studied the binding properties between the human receptor ACE2 (hACE2) and the VOC RBDs and resolved the crystal and cryoelectron microscopy structures of the omicron RBD-hACE2 complex as well as the crystal structure of the delta RBD-hACE2 complex. We found that, unlike alpha, beta, and gamma, omicron RBD binds to hACE2 at a similar affinity to that of the prototype RBD, which might be due to compensation of multiple mutations for both immune escape and transmissibility. The complex structures of omicron RBD-hACE2 and delta RBD-hACE2 reveal the structural basis of how RBD-specific mutations bind to hACE2.
Assuntos
Enzima de Conversão de Angiotensina 2/química , Receptores Virais/química , SARS-CoV-2/química , Sequência de Aminoácidos , Microscopia Crioeletrônica , Humanos , Modelos Moleculares , Mutação/genética , Filogenia , Ligação Proteica , Domínios Proteicos , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/genética , Glicoproteína da Espícula de Coronavírus/ultraestrutura , Eletricidade Estática , Homologia Estrutural de ProteínaRESUMO
Non-segmented negative-strand RNA viruses, including Ebola virus (EBOV), rabies virus, human respiratory syncytial virus and pneumoviruses, can cause respiratory infections, haemorrhagic fever and encephalitis in humans and animals, and are considered a substantial health and economic burden worldwide1. Replication and transcription of the viral genome are executed by the large (L) polymerase, which is a promising target for the development of antiviral drugs. Here, using the L polymerase of EBOV as a representative, we show that de novo replication of L polymerase is controlled by the specific 3' leader sequence of the EBOV genome in an enzymatic assay, and that formation of at least three base pairs can effectively drive the elongation process of RNA synthesis independent of the specific RNA sequence. We present the high-resolution structures of the EBOV L-VP35-RNA complex and show that the 3' leader RNA binds in the template entry channel with a distinctive stable bend conformation. Using mutagenesis assays, we confirm that the bend conformation of the RNA is required for the de novo replication activity and reveal the key residues of the L protein that stabilize the RNA conformation. These findings provide a new mechanistic understanding of RNA synthesis for polymerases of non-segmented negative-strand RNA viruses, and reveal important targets for the development of antiviral drugs.
Assuntos
Ebolavirus , RNA Viral , RNA Polimerase Dependente de RNA , Replicação Viral , Animais , Humanos , Antivirais/farmacologia , Ebolavirus/enzimologia , Ebolavirus/genética , Ebolavirus/crescimento & desenvolvimento , Doença pelo Vírus Ebola/virologia , RNA Viral/biossíntese , RNA Viral/química , RNA Viral/genética , RNA Viral/metabolismo , RNA Polimerase Dependente de RNA/antagonistas & inibidores , RNA Polimerase Dependente de RNA/metabolismo , Genoma Viral , Conformação de Ácido Nucleico , Mutagênese , Estabilidade de RNARESUMO
Filoviruses, including Ebola virus, pose an increasing threat to the public health. Although two therapeutic monoclonal antibodies have been approved to treat the Ebola virus disease1,2, there are no approved broadly reactive drugs to control diverse filovirus infection. Filovirus has a large polymerase (L) protein and the cofactor viral protein 35 (VP35), which constitute the basic functional unit responsible for virus genome RNA synthesis3. Owing to its conservation, the L-VP35 polymerase complex is a promising target for broadly reactive antiviral drugs. Here we determined the structure of Ebola virus L protein in complex with tetrameric VP35 using cryo-electron microscopy (state 1). Structural analysis revealed that Ebola virus L possesses a filovirus-specific insertion element that is essential for RNA synthesis, and that VP35 interacts extensively with the N-terminal region of L by three protomers of the VP35 tetramer. Notably, we captured the complex structure in a second conformation with the unambiguous priming loop and supporting helix away from polymerase active site (state 2). Moreover, we demonstrated that the century-old drug suramin could inhibit the activity of the Ebola virus polymerase in an enzymatic assay. The structure of the L-VP35-suramin complex reveals that suramin can bind at the highly conserved NTP entry channel to prevent substrates from entering the active site. These findings reveal the mechanism of Ebola virus replication and may guide the development of more powerful anti-filovirus drugs.
Assuntos
Microscopia Crioeletrônica , RNA Polimerases Dirigidas por DNA , Ebolavirus , Proteínas Virais Reguladoras e Acessórias , Antivirais/farmacologia , Domínio Catalítico , RNA Polimerases Dirigidas por DNA/química , RNA Polimerases Dirigidas por DNA/metabolismo , RNA Polimerases Dirigidas por DNA/ultraestrutura , Ebolavirus/enzimologia , Doença pelo Vírus Ebola/tratamento farmacológico , Doença pelo Vírus Ebola/virologia , Humanos , Subunidades Proteicas/química , Subunidades Proteicas/metabolismo , RNA Viral/biossíntese , Suramina/química , Suramina/metabolismo , Suramina/farmacologia , Suramina/uso terapêutico , Proteínas Virais Reguladoras e Acessórias/química , Proteínas Virais Reguladoras e Acessórias/metabolismo , Proteínas Virais Reguladoras e Acessórias/ultraestrutura , Replicação ViralRESUMO
Genome transcription and replication of influenza A virus (FluA), catalyzed by viral RNA polymerase (FluAPol), are delicately controlled across the virus life cycle. A switch from transcription to replication occurring at later stage of an infection is critical for progeny virion production and viral non-structural protein NS2 has been implicated in regulating the switch. However, the underlying regulatory mechanisms and the structure of NS2 remained elusive for years. Here, we determine the cryo-EM structure of the FluAPol-NS2 complex at ~3.0 Å resolution. Surprisingly, three domain-swapped NS2 dimers arrange three symmetrical FluPol dimers into a highly ordered barrel-like hexamer. Further structural and functional analyses demonstrate that NS2 binding not only hampers the interaction between FluAPol and the Pol II CTD because of steric conflicts, but also impairs FluAPol transcriptase activity by stalling it in the replicase conformation. Moreover, this is the first visualization of the full-length NS2 structure. Our findings uncover key molecular mechanisms of the FluA transcription-replication switch and have implications for the development of antivirals.
RESUMO
Arenaviruses can cause severe haemorrhagic fever and neurological diseases in humans and other animals, exemplified by Lassa mammarenavirus, Machupo mammarenavirus and lymphocytic choriomeningitis virus, posing great threats to public health1-4. These viruses encode a large multi-domain RNA-dependent RNA polymerase for transcription and replication of the viral genome5. Viral polymerases are one of the leading antiviral therapeutic targets. However, the structure of arenavirus polymerase is not yet known. Here we report the near-atomic resolution structures of Lassa and Machupo virus polymerases in both apo and promoter-bound forms. These structures display a similar overall architecture to influenza virus and bunyavirus polymerases but possess unique local features, including an arenavirus-specific insertion domain that regulates the polymerase activity. Notably, the ordered active site of arenavirus polymerase is inherently switched on, without the requirement for allosteric activation by 5'-viral RNA, which is a necessity for both influenza virus and bunyavirus polymerases6,7. Moreover, dimerization could facilitate the polymerase activity. These findings advance our understanding of the mechanism of arenavirus replication and provide an important basis for developing antiviral therapeutics.
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Arenavirus do Novo Mundo/enzimologia , Microscopia Crioeletrônica , Vírus Lassa/enzimologia , RNA Polimerase Dependente de RNA/química , RNA Polimerase Dependente de RNA/ultraestrutura , Replicação Viral , Apoenzimas/química , Apoenzimas/metabolismo , Apoenzimas/ultraestrutura , Arenavirus do Novo Mundo/ultraestrutura , Domínio Catalítico , Vírus Lassa/ultraestrutura , Vírus da Coriomeningite Linfocítica/enzimologia , Vírus da Coriomeningite Linfocítica/ultraestrutura , Modelos Moleculares , Regiões Promotoras Genéticas/genética , RNA Polimerase Dependente de RNA/metabolismoRESUMO
CRISPR-Cas systems are widespread adaptive antiviral systems used in prokaryotes. Some phages, in turn, although have small genomes can economize the use of genetic space to encode compact or incomplete CRISPR-Cas systems to inhibit the host and establish infection. Phage ICP1, infecting Vibrio cholerae, encodes a compact type I-F CRISPR-Cas system to suppress the antiphage mobile genetic element in the host genome. However, the mechanism by which this compact system recognizes the target DNA and executes interference remains elusive. Here, we present the electron cryo-microscopy (cryo-EM) structures of both apo- and DNA-bound ICP1 surveillance complexes (Aka Csy complex). Unlike most other type I surveillance complexes, the ICP1 Csy complex lacks the Cas11 subunit or a structurally homologous domain, which is crucial for dsDNA binding and Cas3 activation in other type I CRISPR-Cas systems. Structural and functional analyses revealed that the compact ICP1 Csy complex alone is inefficient in binding to dsDNA targets, presumably stalled at a partial R-loop conformation. The presence of Cas2/3 facilitates dsDNA binding and allows effective dsDNA target cleavage. Additionally, we found that Pseudomonas aeruginosa Cas2/3 efficiently cleaved the dsDNA target presented by the ICP1 Csy complex, but not vice versa. These findings suggest a unique mechanism for target dsDNA binding and cleavage by the compact phage-derived CRISPR-Cas system.
Assuntos
Bacteriófagos , Proteínas Associadas a CRISPR , Bacteriófagos/genética , Sistemas CRISPR-Cas , DNA , Proteínas Associadas a CRISPR/metabolismoRESUMO
African swine fever (ASF) is a highly contagious viral disease that affects domestic and wild pigs. The causative agent of ASF is African swine fever virus (ASFV), a large double-stranded DNA virus with a complex virion structure. Among the various proteins encoded by ASFV, A137R is a crucial structural protein associated with its virulence. However, the structure and molecular mechanisms underlying the functions of A137R remain largely unknown. In this study, we present the structure of A137R determined by cryogenic electron microscopy single-particle reconstruction, which reveals that A137R self-oligomerizes to form a dodecahedron-shaped cage composed of 60 polymers. The dodecahedron is literally equivalent to a T = 1 icosahedron where the icosahedral vertexes are located in the center of each dodecahedral facet. Within each facet, five A137R protomers are arranged in a head-to-tail orientation with a long N-terminal helix forming the edge through which adjacent facets stitch together to form the dodecahedral cage. Combining structural analysis and biochemical evidence, we demonstrate that the N-terminal domain of A137R is crucial and sufficient for mediating the assembly of the dodecahedron. These findings imply the role of A137R cage as a core component in the icosahedral ASFV virion and suggest a promising molecular scaffold for nanotechnology applications. IMPORTANCE: African swine fever (ASF) is a lethal viral disease of pigs caused by African swine fever virus (ASFV). No commercial vaccines and antiviral treatments are available for the prevention and control of the disease. A137R is a structural protein of ASFV that is associated with its virulence. The discovery of the dodecahedron-shaped cage structure of A137R in this study is of great importance in understanding ASFV pathogenicity. This finding sheds light on the molecular mechanisms underlying the functions of A137R. Furthermore, the dodecahedral cage formed by A137R shows promise as a molecular scaffold for nanoparticle vectors. Overall, this study provides valuable insights into the structure and function of A137R, contributing to our understanding of ASFV and potentially opening up new avenues for the development of vaccines or treatments for ASF.
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Vírus da Febre Suína Africana , Suínos , Proteínas Estruturais Virais , Animais , Febre Suína Africana/virologia , Vírus da Febre Suína Africana/química , Vírus da Febre Suína Africana/crescimento & desenvolvimento , Vírus da Febre Suína Africana/patogenicidade , Vírus da Febre Suína Africana/ultraestrutura , Microscopia Crioeletrônica , Relação Estrutura-Atividade , Suínos/virologia , Proteínas Estruturais Virais/química , Proteínas Estruturais Virais/metabolismo , Proteínas Estruturais Virais/ultraestrutura , Vírion/química , Vírion/metabolismo , Vírion/ultraestrutura , VirulênciaRESUMO
Swine coronavirus-porcine epidemic diarrhea virus (PEDV) with specific susceptibility to pigs has existed for decades, and recurrent epidemics caused by mutant strains have swept the world again since 2010. In this study, single-cell RNA sequencing was used to perform for the first time, to our knowledge, a systematic analysis of pig jejunum infected with PEDV. Pig intestinal cell types were identified by representative markers and identified a new tuft cell marker, DNAH11. Excepting enterocyte cells, the goblet and tuft cells confirmed susceptibility to PEDV. Enrichment analyses showed that PEDV infection resulted in upregulation of cell apoptosis, junctions, and the MAPK signaling pathway and downregulation of oxidative phosphorylation in intestinal epithelial cell types. The T cell differentiation and IgA production were decreased in T and B cells, respectively. Cytokine gene analyses revealed that PEDV infection downregulated CXCL8, CXCL16, and IL34 in tuft cells and upregulated IL22 in Th17 cells. Further studies found that infection of goblet cells with PEDV decreased the expression of MUC2, as well as other mucin components. Moreover, the antimicrobial peptide REG3G was obviously upregulated through the IL33-STAT3 signaling pathway in enterocyte cells in the PEDV-infected group, and REG3G inhibited the PEDV replication. Finally, enterocyte cells expressed almost all coronavirus entry factors, and PEDV infection caused significant upregulation of the coronavirus receptor ACE2 in enterocyte cells. In summary, this study systematically investigated the responses of different cell types in the jejunum of piglets after PEDV infection, which deepened the understanding of viral pathogenesis.
Assuntos
Infecções por Coronavirus , Vírus da Diarreia Epidêmica Suína , Suínos , Animais , Vírus da Diarreia Epidêmica Suína/genética , Transcriptoma , Intestino Delgado/patologia , Intestinos/patologia , Análise de Sequência de RNARESUMO
The C-C motif chemokine receptor 8 (CCR8) is a class A G-protein-coupled receptor that has emerged as a promising therapeutic target in cancer and autoimmune diseases. In the present study, we solved the cryo-electron microscopy (cryo-EM) structure of the human CCR8-Gi complex in the absence of a ligand at 2.58 Å. Structural analysis and comparison revealed that our apo CCR8 structure undergoes some conformational changes and is similar to that in the CCL1-CCR8 complex structure, indicating an active state. In addition, the key residues of CCR8 involved in the recognition of LMD-009, a potent nonpeptide agonist, were investigated by mutating CCR8 and testing the calcium flux induced by LMD-009-CCR8 interaction. Three mutants of CCR8, Y1133.32A, Y1724.64A, and E2867.39A, showed a dramatically decreased ability in mediating calcium mobilization, indicating their key interaction with LMD-009 and key roles in activation. These structural and biochemical analyses enrich molecular insights into the agonism and activation of CCR8 and will facilitate CCR8-targeted therapy.
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Microscopia Crioeletrônica , Receptores CCR8 , Humanos , Receptores CCR8/metabolismo , Receptores CCR8/química , Receptores CCR8/genética , Modelos Moleculares , Conformação Proteica , Cálcio/metabolismo , Células HEK293RESUMO
BACKGROUND: People with human immunodeficiency virus (HIV) (PWH) have an increased risk of cardiovascular disease (CVD). Cardiac magnetic resonance (CMR) has documented higher myocardial fibrosis, inflammation, and steatosis in PWH, but studies have mostly relied on healthy volunteers as comparators and focused on men. METHODS: We investigated the associations of HIV and HIV-specific factors with CMR phenotypes in female participants enrolled in the Women's Interagency HIV Study's New York and San Francisco sites. Primary phenotypes included myocardial native (n) T1 (fibro-inflammation), extracellular volume fraction (fibrosis), and triglyceride content (steatosis). Associations were evaluated with multivariable linear regression, and results pooled or meta-analyzed across centers. RESULTS: Among 261 women with HIV (WWH, N = 362), 76.2% had undetectable viremia at CMR. For the 82.8% receiving continuous antiretroviral therapy (ART) in the preceding 5 years, adherence was 51.7%, and 69.4% failed to achieve persistent viral suppression (40.7% with peak viral load <200â cp/mL). Overall, WWH showed higher nT1 than women without HIV after full adjustment. This higher nT1 was more pronounced in those with antecedent or current viremia or nadir CD4+ count <200â cells/µL, with the latter also associated with higher extracellular volume fraction. WWH and current CD4+ count <200â cells/µL had less cardiomyocyte steatosis. Cumulative exposure to specific ART showed no associations. CONCLUSIONS: Compared with sociodemographically similar women without HIV, WWH on ART exhibit higher myocardial fibro-inflammation, which is more prominent with unsuppressed viremia or CD4+ lymphopenia. These findings support the importance of improved ART adherence strategies, along with better understanding of latent infection, to mitigate cardiac end-organ damage in this population.
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Infecções por HIV , Humanos , Feminino , Infecções por HIV/tratamento farmacológico , Infecções por HIV/complicações , Pessoa de Meia-Idade , Adulto , Cardiomiopatias , Carga Viral , Fatores de Risco , Imageamento por Ressonância Magnética , Miocárdio/patologiaRESUMO
Microbial remediation of heavy metal polluted environment is ecofriendly and cost effective. Therefore, in the present study, Shewanella putrefaciens stain 4H was previously isolated by our group from the activated sludge of secondary sedimentation tank in a dyeing wastewater treatment plant. The bacterium was able to reduce chromate effectively. The strains showed significant ability to reduce Cr(VI) in the pH range of 8.0 to 10.0 (optimum pH 9.0) and 25-42 â (optimum 30 â) and were able to reduce 300 mg/L of Cr(VI) in 72 h under parthenogenetic anaerobic conditions. In this paper, the complete genome sequence was obtained by Nanopore sequencing technology and analyzed chromium metabolism-related genes by comparative genomics The genomic sequence of S. putrefaciens 4H has a length of 4,631,110 bp with a G + C content of 44.66% and contains 4015 protein-coding genes and 3223, 2414, 2343 genes were correspondingly annotated into the COG, KEGG, and GO databases. The qRT-PCR analysis showed that the expression of chrA, mtrC, and undA genes was up-regulated under Cr(VI) stress. This study explores the Chromium Metabolism-Related Genes of S. putrefaciens 4H and will help to deepen our understanding of the mechanisms of Cr(VI) tolerance and reduction in this strain, thus contributing to the better application of S. putrefaciens 4H in the field of remediation of chromium-contaminated environments.
Assuntos
Shewanella putrefaciens , Shewanella putrefaciens/genética , Shewanella putrefaciens/metabolismo , Oxirredução , Cromo/toxicidade , Cromo/metabolismo , Bactérias/metabolismoRESUMO
At the 3' end of the C2 gene in the mammalian TRB locus, a distinct reverse TRBV30 gene (named TRBV31 in mice) has been conserved throughout evolution. In the fully annotated TRB locus of 14 mammals (including six orders), we observed noteworthy variations in the localization and quality of the reverse V30 genes and Recombination Signal Sequences (RSSs) in the gene trees of 13 mammals. Conversely, the forward V29 genes and RSSs were generally consistent with the species tree of their corresponding species. This finding suggested that the evolution of the reverse V30 gene was not synchronous and likely played a crucial role in regulating adaptive immune responses. To further investigate this possibility, we utilized single-cell TCR sequencing (scTCR-seq) and high-throughput sequencing (HTS) to analyze TCRß CDR3 repertoires from both central and peripheral tissues of Primates (Homo sapiens and Macaca mulatta), Rodentia (Mus musculus: BALB/c, C57BL/6, and Kunming mice), Artiodactyla (Bos taurus and Bubalus bubalis), and Chiroptera (Rhinolophus affinis and Hipposideros armige). Our investigation revealed several novel observations: (1) The reverse V30 gene exhibits classical rearrangement patterns adhering to the '12/23 rule' and the 'D-J rearrangement preceding the V-(D-J) rearrangement'. This results in the formation of rearranged V30-D2J2, V30-D1J1, and V30-D1J2. However, we also identified 'special rearrangement patterns' wherein V30-D rearrangement preceding D-J rearrangement, giving rise to rearranged V30-D2-J1 and forward Vx-D2-J. (2) Compared to the 'deletional rearrangement' (looping out) of forward V1-V29 genes, the reverse V30 gene exhibits preferential utilization with 'inversional rearrangement'. This may be attributed to the shorter distance between the V30 gene and D gene and the 'inversional rearrangement' modes. In summary, in the mammalian TRB locus, the reverse V30 gene has been uniquely preserved throughout evolution and preferentially utilized in V(D)J recombination, potentially serving a significant role in adaptive immunity. These results will pave the way for novel and specialized research into the mechanisms, efficiency, and function of V(D)J recombination in mammals.
Assuntos
Evolução Molecular , Mamíferos , Animais , Mamíferos/genética , Humanos , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Filogenia , Sequenciamento de Nucleotídeos em Larga Escala , CamundongosRESUMO
BACKGROUND: Marginal zone lymphomas (MZLs) comprise a diverse group of indolent lymphoproliferative disorders; however, some patients develop histologic transformation (HT) with rapid progression to aggressive lymphoma. METHODS: Forty-three MZLs with HT (HT-MZLs), 535 MZLs, and 174 de novo diffuse large B-cell lymphomas (DLBCLs) without rearrangements of MYC, BCL2, and BCL6 were collected. Among these, 22 HT-MZLs, 39 MZLs, and 174 DLBCLs were subjected to 148-gene targeted exome sequencing. The clinicopathologic features of patients who had HT-MZL and their genetic alterations were compared with those of patients who had MZLs and DLBCLs. RESULTS: All 43 HT-MZLs corresponded to DLBCLs. No HT-MZLs harbored BCL2 and MYC and/or BCL6 rearrangements. Bone marrow involvement and higher levels of lactate dehydrogenase were significantly more common in HT-MZLs than in MZLs. Furthermore, upregulated BCL6, MUM1, C-MYC, and Ki-67 expression was observed more frequently in HT-MZLs than in MZLs. TBL1XR1 was the most frequently altered gene (63.6%) in HT-MZLs, followed by CCND3 (31.8%), CARD11, ID3, and TP53 (22.7%). A trend toward worse progression-free survival in patients with TBL1XR1 mutations was observed. Compared with MZLs and non-germinal center B-cell (GCB) type DLBCLs, significantly higher frequencies of TBL1XR1 and ID3 mutations were identified in HT-MZLs. PIM1 mutations frequently occurred in DLBCLs and were significantly associated with TBL1XR1 mutations but were mutated less in HT-MZLs that had TBL1XR1 mutations. CONCLUSIONS: The current findings reveal the clinicopathologic and genetic features of HT-MZLs, suggesting that these tumors might constitute a group distinct from MZL and de novo non-GCB type DLBCL. TBL1XR1 mutations may be considered a predictor of HT in MZL.
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Linfoma de Zona Marginal Tipo Células B , Linfoma Difuso de Grandes Células B , Humanos , Linfoma de Zona Marginal Tipo Células B/genética , Linfoma Difuso de Grandes Células B/genética , Linfoma Difuso de Grandes Células B/patologia , Intervalo Livre de Progressão , Proteínas Proto-Oncogênicas c-bcl-2/genéticaRESUMO
Branch angle (BA) is a critical morphological trait that significantly influences planting density, light interception and ultimately yield in plants. Despite its importance, the regulatory mechanism governing BA in rapeseed remains poorly understood. In this study, we generated 109 transcriptome data sets for 37 rapeseed accessions with divergent BA phenotypes. Relative to adaxial branch segments, abaxial segments accumulated higher levels of auxin and exhibited lower expression of six TCP1 homologues and one GA20ox3. A co-expression network analysis identified two modules highly correlated with BA. The modules contained homologues to known BA control genes, such as FUL, YUCCA6, TCP1 and SGR3. Notably, a homoeologous exchange (HE), occurring at the telomeres of A09, was prevalent in large BA accessions, while an A02-C02 HE was common in small BA accessions. In their corresponding regions, these HEs explained the formation of hub gene hotspots in the two modules. QTL-seq analysis confirmed that the presence of a large A07-C06 HE (~8.1 Mb) was also associated with a small BA phenotype, and BnaA07.WRKY40.b within it was predicted as candidate gene. Overexpressing BnaA07.WRKY40.b in rapeseed increased BA by up to 20°, while RNAi- and CRISPR-mediated mutants (BnaA07.WRKY40.b and BnaC06.WRKY40.b) exhibited decreased BA by up to 11.4°. BnaA07.WRKY40.b was exclusively localized to the nucleus and exhibited strong expression correlations with many genes related to gravitropism and plant architecture. Taken together, our study highlights the influence of HEs on rapeseed plant architecture and confirms the role of WRKY40 homologues as novel regulators of BA.
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Locos de Características Quantitativas , Transcriptoma , Transcriptoma/genética , Locos de Características Quantitativas/genética , Brassica rapa/genética , Regulação da Expressão Gênica de Plantas , Brassica napus/genética , Brassica napus/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Ácidos Indolacéticos/metabolismo , Fenótipo , Genes de Plantas/genéticaRESUMO
IMPORTANCE: Porcine epidemic diarrhea (PED) caused by PED virus (PEDV) remains a big threat to the swine industry worldwide. Vaccination with live attenuated vaccine is a promising method to prevent and control PED, because it can elicit a more protective immunity than the killed vaccine, subunit vaccine, and so on. In this study, we found two obvious deletions in the genome of a high passage of AH2012/12. We further confirmed the second deletion which contains seven amino acids at the carboxy-terminus of the S2 gene and the start codon of ORF3 can reduce its pathogenicity in vivo. Animal experiments indicated that the recombinant PEDV with deleted carboxy-terminus of S gene showed higher IgG, IgA, neutralization antibodies, and protection effects against virus challenge than the killed vaccine. These data reveal that the engineering of the carboxy-terminus of the S2 gene may be a promising method to develop live attenuated vaccine candidates of PEDV.
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Infecções por Coronavirus , Vírus da Diarreia Epidêmica Suína , Doenças dos Suínos , Animais , Infecções por Coronavirus/veterinária , Infecções por Coronavirus/virologia , Diarreia , Vírus da Diarreia Epidêmica Suína/genética , Vírus da Diarreia Epidêmica Suína/patogenicidade , Suínos , Doenças dos Suínos/virologia , Vacinas Atenuadas/genética , Vacinas de Produtos Inativados , Vacinas Virais/genética , VirulênciaRESUMO
IMPORTANCE: As an emerging porcine enteropathogenic coronavirus that has the potential to infect humans, porcine deltacoronavirus (PDCoV) is receiving increasing attention. However, no effective commercially available vaccines against this virus are available. In this work, we designed a spike (S) protein and receptor-binding domain (RBD) trimer as a candidate PDCoV subunit vaccine. We demonstrated that S protein induced more robust humoral and cellular immune responses than the RBD trimer in mice. Furthermore, the protective efficacy of the S protein was compared with that of inactivated PDCoV vaccines in piglets and sows. Of note, the immunized piglets and suckling pig showed a high level of NAbs and were associated with reduced virus shedding and mild diarrhea, and the high level of NAbs was maintained for at least 4 months. Importantly, we demonstrated that S protein-based subunit vaccines conferred significant protection against PDCoV infection.
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Infecções por Coronavirus , Coronavirus , Doenças dos Suínos , Vacinas de Subunidades Antigênicas , Animais , Feminino , Humanos , Camundongos , Coronavirus/genética , Infecções por Coronavirus/prevenção & controle , Infecções por Coronavirus/veterinária , Deltacoronavirus , Suínos , Vacinas de Subunidades Antigênicas/administração & dosagemRESUMO
In the face of rising global temperatures, the mechanisms behind an organism's ability to acclimate to heat stress remain enigmatic. The rice leaf folder, Cnaphalocrocis medinalis, traditionally viewed as temperature-sensitive, paradoxically exhibits robust larval acclimation to heat stress. This study used the heat-acclimated strain HA39, developed through multigenerational exposure to 39°C during the larval stage, and the unacclimated strain HA27 reared at 27°C to unravel the transgenerational effects of heat acclimation and its regulatory mechanisms. Heat acclimation for larvae incurred a fitness cost in pupae when exposed to high temperature, yet a significant transgenerational effect surfaced, revealing heightened fitness benefit in pupae from HA39, even without additional heat exposure during larval recovery at 27°C. This transgenerational effect exhibited a short-term memory, diminishing after two recovery generations. Moreover, the effect correlated with increased superoxide dismutase (SOD) enzyme activity and expression levels of oxidoreductase genes, representing physiological and molecular foundations of heat acclimation. Heat-acclimated larvae displayed elevated DNA methylation levels, while pupae from HA39, in recovery generations, exhibited decreased methylation indicated by the upregulation of a demethylase gene and downregulation of two methyltransferase genes at high temperatures. In summary, heat acclimation induces DNA methylation, orchestrating heat-stress memory and influencing the expression levels of oxidoreductase genes and SOD activity. Heat-stress memory enhances the acclimation of the migratory insect pest to global warming.
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This study aimed to optimize the sampling of spin-lock times (TSLs) in quantitative T1ρ mapping for improved reproducibility. Two new TSL sampling schemes were proposed: (i) reproducibility-guided random sampling (RRS) and (ii) reproducibility-guided optimal sampling (ROS). They were compared to the existing linear sampling (LS) and precision-guided sampling (PS) schemes for T1ρ reproducibility through numerical simulations, phantom experiments, and volunteer studies. Each study evaluated the four sampling schemes with three commonly used T1ρ preparations based on composite and balanced spin-locking. Additionally, the phantom and volunteer studies investigated the impact of B0 and B1 field inhomogeneities on T1ρ reproducibility, respectively. The reproducibility was assessed using the coefficient of variation (CoV) by repeating the T1ρ measurements eight times for phantom experiments and five times for volunteer studies. Numerical simulations resulted in lower mean CoVs for the proposed RRS (1.74%) and ROS (0.68%) compared to LS (2.93%) and PS (3.68%). In the phantom study, the mean CoVs were also lower for RRS (2.7%) and ROS (2.6%) compared to LS (4.1%) and PS (3.1%). Furthermore, the mean CoVs of the proposed RRS and ROS were statistically lower (P < 0.001) compared to existing LS and PS schemes at a B1 offset of 20%. In the volunteer study, consistently lower mean CoVs were observed in bilateral thigh muscles for RRS (9.3%) and ROS (9.2%) compared to LS (10.9%) and PS (10.2%), and the difference was more prominent at B0 offsets higher than 50 Hz. The proposed sampling schemes improve the reproducibility of quantitative T1ρ mapping by optimizing the selection of TSLs. This improvement is especially beneficial for longitudinal studies that track and monitor disease progression and treatment response.
RESUMO
Diabetes mellitus is associated with a wide range of neuropathies, vasculopathies, and immunopathies, resulting in many complications. More than 30% of diabetic patients risk developing diabetic foot ulcers (DFUs). Non-coding RNAs (ncRNAs), including microRNAs (miRNAs), long non-coding RNAs (lncRNAs), and circular RNAs (circRNAs), play essential roles in various biological functions in the hyperglycaemic environment that determines the development of DFU. Ulceration results in tissue breakdown and skin barrier scavenging, thereby facilitating bacterial infection and biofilm formation. Many bacteria contribute to diabetic foot infection (DFI), including Staphylococcus aureus (S. aureus) et al. A heterogeneous group of "ncRNAs," termed small RNAs (sRNAs), powerfully regulates biofilm formation and DFI healing. Multidisciplinary foot care interventions have been identified for nonhealing ulcers. With an appreciation of the link between disease processes and ncRNAs, a novel therapeutic model of bioactive materials loaded with ncRNAs has been developed to prevent and manage diabetic foot complications.
Assuntos
Infecções Bacterianas , Diabetes Mellitus , Pé Diabético , Humanos , Pé Diabético/genética , Pé Diabético/terapia , Pé Diabético/complicações , Staphylococcus aureus , Infecções Bacterianas/complicações , Pé , Cicatrização/genéticaRESUMO
Circular RNAs (circRNAs), a class of non-coding RNA molecules, are recognized for their unique functions; however, their responses to herbicide stress in Brassica napus remain unclear. In this study, the role of circRNAs in response to herbicide treatment was investigated in two rapeseed cultivars: MH33, which confers non-target-site resistance (NTSR), and EM28, which exhibits target-site resistance (TSR). The genome-wide circRNA profiles of herbicide-stressed and non-stressed seedlings were analyzed. The findings indicate that NTSR seedlings exhibited a greater abundance of circRNAs, shorter lengths of circRNAs and their parent genes, and more diverse functions of parent genes compared with TSR seedlings. Compared to normal-growth plants, the herbicide-stressed group exhibited similar trends in the number of circRNAs, functions of parent genes, and differentially expressed circRNAs as observed in NTSR seedlings. In addition, a greater number of circRNAs that function as competing microRNA (miRNA) sponges were identified in the herbicide stress and NTSR groups compared to the normal-growth and TSR groups, respectively. The differentially expressed circRNAs were validated by qPCR. The differntially expressed circRNA-miRNA networks were predicted, and the mRNAs targeted by these miRNAs were annotated. Our results suggest that circRNAs play a crucial role in responding to herbicide stress, exhibiting distinct responses between NTSR and TSR in rapeseed. These findings offer valuable insights into the mechanisms underlying herbicide resistance in rapeseed.