RESUMO
NACs (NAMãATAF1/2ãCUC1/2), as a large family of plant transcription factors, are widely involved in abiotic stress responses. This study aimed to isolate and clone a novel stress-responsive transcription factor LpNAC5 from Lilium pumilum bulbs. Drought, salt, alkali, and ABA stresses induced the expression of LpNAC5. Transgenic tobacco plants overexpressing LpNAC5 were generated using the Agrobacterium-mediated method to understand the role of this factor in stress response. These plants exhibited increased tolerance to drought, salt, and alkali stresses. The tobacco plants overexpressing LpNAC5 showed strong drought, salt, and alkaline tolerance. Under the three abiotic stresses, the activities of antioxidant enzymes were enhanced, the contents of proline and chlorophyll increased, and the contents of malondialdehyde decreased. The functional analysis revealed that LpNAC5 enabled plants to positively regulate drought and salt stresses. These findings not only provided valuable insights into stress tolerance mechanisms in L. pumilum but also offered a potential genetic resource for breedi.
Assuntos
Secas , Regulação da Expressão Gênica de Plantas , Lilium , Proteínas de Plantas , Plantas Geneticamente Modificadas , Tolerância ao Sal , Estresse Fisiológico , Lilium/genética , Lilium/metabolismo , Plantas Geneticamente Modificadas/genética , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Tolerância ao Sal/genética , Estresse Fisiológico/genética , Nicotiana/genética , Nicotiana/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Estresse SalinoRESUMO
Subgroup J avian leukosis virus (ALV-J) is a major pathogen in poultry, causing substantial economic losses to the poultry industry worldwide. Exosomal small RNAs derived from virus-infected cells or biological fluids can serve as viral transmission vectors. However, the role and mechanism of exosomal miRNA in ALV-J infection are unclear. In this study, we demonstrated that exosomal microRNA-7-25207 (miR-7-25207) could increase the titers of ALV-J. Exosomes isolated from ALV-J-infected DF-1 cells (Exo-ALV-J) contained partial viral proteins from ALV-J and could transmit the infection to uninfected DF-1 cells, leading to productive infection. Additionally, the RNA expression profile of exosomes was altered following ALV-J infection. miRNA analysis revealed that the expression of exosomal miR-7-25207 increased. Overexpression of miR-7-25207 significantly increased the titers of ALV-J in transfected cells. Furthermore, miR-7-25207 directly suppressed the expression of Akt and PRC1. Akt, in turn, directly inhibited CyclinQ1 expression, while PRC1 directly interfered with YAF2 expression. In conclusion, ALV-J infection activates the expression of miR-7-25207, which is subsequently delivered via exosomes to uninfected cells, increasing ALV-J titers by targeting Akt-CyclinQ1 and PRC1-YAF2 dual pathways. These findings suggest that exosomal miR-7-25207 may serve as a potential biomarker for clinical parameters in ALV-J infection.
RESUMO
PURPOSE: In this study, the effect of heparin-derived oligosaccharide (HDO) on vascular endothelial growth factor (VEGF) induced vascular smooth muscle cell (VSMC) proliferation and the signal transduction mechanisms involved were investigated. METHODS: MTT assays were used to measure VSMC proliferation, flow cytometry to analyze cell cycle distribution, RT-PCR for detection of gene transcript levels, and cell-based ELISA, Western blotting and immunocytochemical methods to detect the expression of PKC-α, ERK 1/2, p-ERK 1/2, Akt, p-Akt, p-PDK1 and p-GSK-3ß. RESULTS: HDO at concentrations of 0.01, 0.1 and 1 µmol·L(-1) dose-dependently inhibited VEGF-induced VSMC proliferation with inhibition indices of 6.8 %, 13.1 % and 28.9 %, respectively. Similar concentrations of HDO dose-dependently decreased the percentage of VEGF-induced cells in S phase to 3.6 %, 3.4 %, and 5.4 %, while increasing that of cells arrested in the G0/G1 phase to 80 %, 82 % and 83.6 %. HDO at 0.01, 0.1 or 1 µmol·L(-1) inhibited VEGF-induced PKC-α mRNA expression, with inhibition indices of 9.2 %, 16.1 % and 54.0 %. HDO at 0.1 or 1 µmol·L(-1) inhibited VEGF-induced proto-oncogene mRNA expression, with inhibition indices of 5.2 % and 6.6 % for c-jun, 8.8 % and 11.6 % for c-myc, and 6.5 % and 11.9 % for c-fos, respectively. Additionally, treatment with 0.01, 0.1 or 1 µmol·L(-1) HDO, inhibited VEGF-induced expression of some proliferation related proteins with inhibition indices of 33.2 %, 56.3 % and 77.0 % for PKC-α, 33.7 %, 38.7 % and 53.2 % for p-Akt, 3.5 %, 24.2 % and 49.3 % for p-ERK 1/2, 39.2 %, 71.8 % and 80.7 % for p-PDK 1 and 41.4 %, 89.4 % and 92.4 % for p-GSK-3ß, respectively. The results showed that HDO inhibited PKC-α, c-jun, c-fos and c-myc mRNA transcription, and also down-regulated phosphorylation levels of ERK 1/2 and Akt. CONCLUSION: Our study demonstrates that HDO inhibits transcription of proliferation-related proto-oncogenes and arrests G1/S transition through inhibition of the PKC, MAPK and Akt/PI3K pathways in association with inhibition of VSMC proliferation. This altered molecular signature may explain one mechanism of HDO-mediated inhibition of VSMC proliferation.
Assuntos
Proliferação de Células/efeitos dos fármacos , Músculo Liso Vascular/metabolismo , Oligossacarídeos/farmacologia , Transdução de Sinais/efeitos dos fármacos , Animais , Western Blotting , Ciclo Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaio de Imunoadsorção Enzimática , Músculo Liso Vascular/citologia , Proto-Oncogenes/efeitos dos fármacos , RNA , Ratos , Fator A de Crescimento do Endotélio Vascular/administração & dosagem , Fator A de Crescimento do Endotélio Vascular/farmacologiaRESUMO
The title compouand {[Zn(C(4)H(4)O(6))(C(12)H(8)N(2))]·6H(2)O}(n), has a linear chain structure parallel to [100] with Zn(C(4)H(4)O(6))(C(12)H(8)N(2)) repeat units; the asymmetric unit consists of one Zn(2+) cation, one l-tartrate dianion, one 1,10-phenanthroline and six free water mol-ecules. The Zn atom is in a distorted octa-hedral ZnN(2)O(4) coordination environment. The crystal structure is stabilized by O-Hâ¯O hydrogen bonds and π-π stacking of the phenanthroline units [centroid-centroid distances in the range 3.552â (2)-3.625â (2)Å] occurs between the chains. The title compound is isotypic with the Cu and Mn analogues.
RESUMO
The title salt, (C(6)H(11)N(2))(2)[Cd(C(7)H(3)NO(4))(2)], displays a discrete mononuclear structure, in which the central Cd(II) atom is six-coordinated in a distorted octa-hedral coordination geometry by two N and four O atoms from two different pyridine-2,6-dicarboxyl-ate anions in an O(2),N,O(6)-tridentate chelation mode. The crystal packing is stabilized by N-Hâ¯O hydrogen bonds and π-π inter-actions [centroid-centroid distance = 3.576â (5)â Å].
RESUMO
In the title compound, [Mn(C(10)H(10)N(2))(6)](ClO(4))(2), the Mn(II) ion, located on an inversion center, is coordinated by six N atoms from three pairs of symmetry-related 1-benzyl-1H-imidazole ligands in a distorted octa-hedral geometry. In the crystal, weak inter-molecular C-Hâ¯O hydrogen bonds link the complex cations and perchlorate anions.
RESUMO
In the title compound, C(14)H(14)N(4), the center of the phenyl-ene group is a crystallographic center of inversion. The compound is composed of three aromatic rings displaying a Z-like conformation. The dihedral angle between the pyrazole rings and the central phenyl ring is 83.84â (9)°.
RESUMO
In the dication of the title compound, C(16)H(20)N(4) (2+)·C(10)H(4)O(8) (2-)·H(2)O, the dihedral angles formed by mean planes of the imidazolium rings and the benzene ring are 69.05â (18) and 89.1â (2)°. In the crystal, the components are linked into a three-dimensional network by inter-molecular N-Hâ¯O and O-Hâ¯O hydrogen bonds.
RESUMO
In the pseudo-centrosymmetric mol-ecule of the title compound, C(18)H(16)N(4), two benzimidazole fragments form the dihedral angles of 83.49â (7) and 79.37â (7)°, with the mean plane of the linking butene chain. No classical inter-molecular inter-actions are observed. The porous crystal packing exhibits voids of 85â Å(3).
RESUMO
The asymmetric unit of the title compound, C(14)H(16)N(4) (2+)·2C(7)H(4)NO(4) (-)·2C(7)H(5)NO(4), comprises one-half of the 3,3'-(p-phenyl-enedimethyl-ene)di-1H-imidazol-1-ium dication, which lies on an inversion centre, one 4-nitro-benzoate anion and one 4-nitro-benzoic acid mol-ecule. In the crystal, the components are linked into a two-dimensional network parallel to (110) by O-Hâ¯O, N-Hâ¯O and C-Hâ¯O hydrogen bonds.
RESUMO
Variations in many genes linked to sporadic Alzheimer's disease (AD) show abundant expression in microglia, but relationships among these genes remain largely elusive. Here, we establish isogenic human ESC-derived microglia-like cell lines (hMGLs) harboring AD variants in CD33, INPP5D, SORL1, and TREM2 loci and curate a comprehensive atlas comprising ATAC-seq, ChIP-seq, RNA-seq, and proteomics datasets. AD-like expression signatures are observed in AD mutant SORL1 and TREM2 hMGLs, while integrative multi-omic analysis of combined epigenetic and expression datasets indicates up-regulation of APOE as a convergent pathogenic node. We also observe cross-regulatory relationships between SORL1 and TREM2, in which SORL1R744X hMGLs induce TREM2 expression to enhance APOE expression. AD-associated SORL1 and TREM2 mutations also impaired hMGL Aß uptake in an APOE-dependent manner in vitro and attenuated Aß uptake/clearance in mouse AD brain xenotransplants. Using this modeling and analysis platform for human microglia, we provide new insight into epistatic interactions in AD genes and demonstrate convergence of microglial AD genes at the APOE locus.
Assuntos
Doença de Alzheimer/genética , Doença de Alzheimer/patologia , Apolipoproteínas E/genética , Variação Genética , Células-Tronco Embrionárias Humanas/metabolismo , Microglia/patologia , Peptídeos beta-Amiloides/metabolismo , Animais , Encéfalo/patologia , Diferenciação Celular , Linhagem Celular , Cromatina/metabolismo , Epigênese Genética , Redes Reguladoras de Genes , Marcação de Genes , Loci Gênicos , Humanos , Camundongos Transgênicos , Modelos Biológicos , Proteínas Mutantes/metabolismo , Mutação/genética , Fagocitose , Proteoma/metabolismo , Transdução de Sinais , Transcriptoma/genética , Transplante Heterólogo , Regulação para Cima/genéticaRESUMO
Mechanisms underlying motor neuron degeneration in amyotrophic lateral sclerosis (ALS) are yet unclear. Specific deletion of the ER-component membralin in astrocytes manifested postnatal motor defects and lethality in mice, causing the accumulation of extracellular glutamate through reducing the glutamate transporter EAAT2. Restoring EAAT2 levels in membralin KO astrocytes limited astrocyte-dependent excitotoxicity in motor neurons. Transcriptomic profiles from mouse astrocytic membralin KO motor cortex indicated significant perturbation in KEGG pathway components related to ALS, including downregulation of Eaat2 and upregulation of Tnfrsf1a. Changes in gene expression with membralin deletion also overlapped with mouse ALS models and reactive astrocytes. Our results shown that activation of TNF receptor (TNFR1)-NFκB pathway known to suppress Eaat2 transcription was upregulated with membralin deletion. Further, reduced membralin and EAAT2 levels correlated with disease progression in spinal cord from SOD1-mutant mouse models, and reductions in membralin/EAAT2 were observed in human ALS spinal cord. Importantly, overexpression of membralin in SOD1G93A astrocytes decreased TNFR1 levels and increased EAAT2 expression, and improved motor neuron survival. Importantly, upregulation of membralin in SOD1G93A mice significantly prolonged mouse survival. Together, our study provided a mechanism for ALS pathogenesis where membralin limited glutamatergic neurotoxicity, suggesting that modulating membralin had potentials in ALS therapy.
Assuntos
Esclerose Lateral Amiotrófica/metabolismo , Astrócitos/metabolismo , Ácido Glutâmico/metabolismo , Córtex Motor/metabolismo , Proteínas do Tecido Nervoso/deficiência , Esclerose Lateral Amiotrófica/genética , Esclerose Lateral Amiotrófica/patologia , Animais , Astrócitos/patologia , Regulação para Baixo , Transportador 2 de Aminoácido Excitatório/biossíntese , Transportador 2 de Aminoácido Excitatório/genética , Ácido Glutâmico/genética , Humanos , Camundongos , Camundongos Knockout , Córtex Motor/patologia , Proteínas do Tecido Nervoso/metabolismo , Receptores Tipo I de Fatores de Necrose Tumoral/biossíntese , Receptores Tipo I de Fatores de Necrose Tumoral/genética , Superóxido Dismutase/biossíntese , Superóxido Dismutase/genética , Transcrição Gênica , Regulação para CimaRESUMO
In the developing brain, immature synapses contain calcium-permeable AMPA glutamate receptors (AMPARs) that are subsequently replaced with GluA2-containing calcium-impermeable AMPARs as synapses stabilize and mature. Here, we show that this essential switch in AMPARs and neuronal synapse maturation is regulated by astrocytes. Using biochemical fractionation of astrocyte-secreted proteins and mass spectrometry, we identified that astrocyte-secreted chordin-like 1 (Chrdl1) is necessary and sufficient to induce mature GluA2-containing synapses to form. This function of Chrdl1 is independent of its role as an antagonist of bone morphogenetic proteins (BMPs). Chrdl1 expression is restricted to cortical astrocytes in vivo, peaking at the time of the AMPAR switch. Chrdl1 knockout (KO) mice display reduced synaptic GluA2 AMPARs, altered kinetics of synaptic events, and enhanced remodeling in an in vivo plasticity assay. Studies have shown that humans with mutations in Chrdl1 display enhanced learning. Thus astrocytes, via the release of Chrdl1, promote GluA2-dependent synapse maturation and thereby limit synaptic plasticity.
Assuntos
Astrócitos/metabolismo , Proteínas do Olho/fisiologia , Proteínas do Tecido Nervoso/fisiologia , Plasticidade Neuronal , Receptores de AMPA/metabolismo , Sinapses/fisiologia , Animais , Células Cultivadas , Proteínas do Olho/metabolismo , Feminino , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas do Tecido Nervoso/metabolismo , Ratos Sprague-DawleyRESUMO
Endoplasmic-reticulum-associated degradation (ERAD) is an important protein quality control system which maintains protein homeostasis. Constituents of the ERAD complex and its role in neurodegeneration are not yet fully understood. Here, using proteomic and FRET analyses, we demonstrate that the ER protein membralin is an ERAD component, which mediates degradation of ER luminal and membrane substrates. Interestingly, we identify nicastrin, a key component of the γ-secretase complex, as a membralin binding protein and membralin-associated ERAD substrate. We demonstrate a reduction of membralin mRNA and protein levels in Alzheimer's disease (AD) brain, the latter of which inversely correlates with nicastrin abundance. Furthermore, membralin deficiency enhances γ-secretase activity and neuronal degeneration. In a mouse AD model, downregulating membralin results in ß-amyloid pathology, neuronal death, and exacerbates synaptic/memory deficits. Our results identify membralin as an ERAD component and demonstrate a critical role for ERAD in AD pathogenesis.
Assuntos
Doença de Alzheimer/patologia , Secretases da Proteína Precursora do Amiloide/metabolismo , Disfunção Cognitiva/patologia , Degradação Associada com o Retículo Endoplasmático/fisiologia , Retículo Endoplasmático/metabolismo , Memória/fisiologia , Proteínas do Tecido Nervoso/metabolismo , Peptídeos beta-Amiloides/metabolismo , Precursor de Proteína beta-Amiloide/metabolismo , Animais , Encéfalo/fisiologia , Linhagem Celular , Feminino , Células HEK293 , Humanos , Masculino , Glicoproteínas de Membrana/metabolismo , Proteínas de Membrana , Camundongos , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas do Tecido Nervoso/genética , Ligação Proteica , Dobramento de Proteína , Interferência de RNA , RNA Mensageiro/genética , RNA Interferente Pequeno/genéticaRESUMO
gp78 is a membrane-anchored ubiquitin ligase mediating the degradation of HMG-CoA reductase (HMGCR) and Insig-1. As a rate-limiting enzyme in cholesterol biosynthesis, HMGCR undergoes rapid sterol-promoted degradation. In contrast, destruction of Insig-1 releases its inhibition on SREBP and stimulates the expression of lipogenic genes. Thus, gp78 has opposite effects on lipid biosynthesis. We here generated liver-specific gp78 knockout (L-gp78(-/-)) mice and showed that although the degradation of HMGCR was blunted, SREBP was suppressed due to the elevation of Insig-1/-2, and therefore the lipid biosynthesis was decreased. The L-gp78(-/-) mice were protected from diet-/age-induced obesity and glucose intolerance. The livers of L-gp78(-/-) mice produced more FGF21, which activated thermogenesis in brown adipocytes and enhanced energy expenditure. Together, the major function of gp78 in liver is regulating lipid biosynthesis through SREBP pathway. Ablation of gp78 decreases the lipid levels and increases FGF21, and is beneficial to patients with metabolic diseases.