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1.
Mol Med ; 26(1): 69, 2020 07 08.
Artigo em Inglês | MEDLINE | ID: mdl-32641037

RESUMO

BACKGROUND: We previously showed that the autophagy inhibitor chloroquine (CQ) increases inflammatory cleaved caspase-1 activity in myocytes, and that caspase-1/11 is protective in sterile liver injury. However, the role of caspase-1/11 in the recovery of muscle from ischemia caused by peripheral arterial disease is unknown. We hypothesized that caspase-1/11 mediates recovery in muscle via effects on autophagy and this is modulated by CQ. METHODS: C57Bl/6 J (WT) and caspase-1/11 double-knockout (KO) mice underwent femoral artery ligation (a model of hind-limb ischemia) with or without CQ (50 mg/kg IP every 2nd day). CQ effects on autophagosome formation, microtubule associated protein 1A/1B-light chain 3 (LC3), and caspase-1 expression was measured using electron microscopy and immunofluorescence. Laser Doppler perfusion imaging documented perfusion every 7 days. After 21 days, in situ physiologic testing in tibialis anterior muscle assessed peak force contraction, and myocyte size and fibrosis was also measured. Muscle satellite cell (MuSC) oxygen consumption rate (OCR) and extracellular acidification rate was measured. Caspase-1 and glycolytic enzyme expression was detected by Western blot. RESULTS: CQ increased autophagosomes, LC3 consolidation, total caspase-1 expression and cleaved caspase-1 in muscle. Perfusion, fibrosis, myofiber regeneration, muscle contraction, MuSC fusion, OCR, ECAR and glycolytic enzyme expression was variably affected by CQ depending on presence of caspase-1/11. CQ decreased perfusion recovery, fibrosis and myofiber size in WT but not caspase-1/11KO mice. CQ diminished peak force in whole muscle, and myocyte fusion in MuSC and these effects were exacerbated in caspase-1/11KO mice. CQ reductions in maximal respiration and ATP production were reduced in caspase-1/11KO mice. Caspase-1/11KO MuSC had significant increases in protein kinase isoforms and aldolase with decreased ECAR. CONCLUSION: Caspase-1/11 signaling affects the response to ischemia in muscle and effects are variably modulated by CQ. This may be critically important for disease treated with CQ and its derivatives, including novel viral diseases (e.g. COVID-19) that are expected to affect patients with comorbidities like cardiovascular disease.


Assuntos
Caspase 1/metabolismo , Caspases Iniciadoras/metabolismo , Cloroquina/farmacologia , Infecções por Coronavirus/patologia , Isquemia/patologia , Músculo Esquelético/patologia , Pneumonia Viral/patologia , Animais , Autofagossomos/metabolismo , Autofagia/efeitos dos fármacos , Betacoronavirus , Infecções por Coronavirus/tratamento farmacológico , Glicólise/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Associadas aos Microtúbulos/metabolismo , Células Musculares/metabolismo , Desenvolvimento Muscular , Músculo Esquelético/metabolismo , Neovascularização Fisiológica , Fosforilação Oxidativa , Pandemias , Doença Arterial Periférica/patologia , Pneumonia Viral/tratamento farmacológico , Regeneração , Transdução de Sinais
2.
Medicine (Baltimore) ; 99(20): e20340, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32443386

RESUMO

BACKGROUND: Esophageal cancer (ESCA) is one of the most deadly malignancies in the world. Although the management and treatment of patients with ESCA have improved, the overall 5-year survival rate is still very poor. METHODS: The study aimed to identify potential key genes associated with the pathogenesis and prognosis of ESCA. In the study, integrated bioinformatics methods were used to screen differentially expressed genes (DEGs) between ESCA and normal tissue in the data set of gene expression profiles. The hub gene in DEGs was further analyzed by protein-protein interaction (PPI) network and survival analysis to explore its relationship with the pathogenesis and poor prognosis of ESCA. RESULTS: 134 up-regulated genes and 183 down-regulated genes were obtained in ESCA compared with normal tissues. Moreover, the PPI network was established with 176 nodes and 800 interactions. Ten hub genes (AURKA, CDC20, BUB1, TOP2A, ASPM, DLGAP5, TPX2, CENPF, UBE2C, and NEK2) were filtered out based on the degree value. Functional enrichment analysis indicated that a variety of extracellular related items and ECM-receptor interaction pathway were all correlated with the ESCA. CONCLUSIONS: The results of this study would provide some guidance for further study of diagnostic and prognostic biomarkers to promote ESCA treatment.


Assuntos
Neoplasias Esofágicas/genética , Mapas de Interação de Proteínas , Biomarcadores Tumorais , Proteínas de Ciclo Celular/metabolismo , Biologia Computacional , Regulação para Baixo , Humanos , Proteínas Associadas aos Microtúbulos/metabolismo , Proteínas Nucleares , Prognóstico , Análise Serial de Proteínas , Transcriptoma , Regulação para Cima
3.
Medicine (Baltimore) ; 99(19): e19980, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32384449

RESUMO

Genome-wide association studies have identified single nucleotide polymorphisms (SNPs) associated with bladder cancer (BCa) risk in Caucasian and East Asian population. The objective of this study was to validate these SNPs in Chinese population and evaluate whether these SNPs could differentiate the individual inherited risk for BCa.A case-control study including 581 BCa cases and 1561 healthy controls was performed. Germline DNA samples from all individuals were genotyped for eight SNPs. Genetic risk score (GRS) was calculated for each individual based on the odds ratios and risk allele frequencies of five risk-associated SNPs.Among eight SNPs evaluated in this study, rs798766 at 4p16.3 [OR = 1.39 (1.15-1.67), P < .001], rs9642880 [OR = 1.17 (1.06-1.30), P < .001] and rs4813953 at 20p12.2 [OR = 1.09 (1.02-1.17), P = .016] were found associated with BCa risk in Chinese population. A genetic risk score was established based on five SNPs (including the above three SNPs and two other SNPs which have the consistent direction with previous reported genome-wide association study). The mean GRS was significantly higher in BCa cases than controls (1.22 vs. 1.01, P < .001). When subjects were categorized into low- (<0.8), average- (0.8-1.2), and high-risk (>1.2) groups, the likelihoods of BCa were 25.2%, 33.7% and 55.0%, respectively (P-trend < 2.2 × 10). In subgroup analyses, no significant difference was observed in mean GRS among BCa patients with different stages or grades.In conclusion, two SNPs derived from East Asian and one SNP from Caucasian were associated with BCa risk in Chinese population. These results provided additional information of genetic risks for BCa in Chinese population. Genetic risk score based on these SNPs can reveal inherited risk of BCa, and may have potential for modifying personalized cancer screening strategy.


Assuntos
Predisposição Genética para Doença/etnologia , Proteínas Associadas aos Microtúbulos/genética , Polimorfismo de Nucleotídeo Único , Receptor Tipo 3 de Fator de Crescimento de Fibroblastos/genética , Neoplasias da Bexiga Urinária , Idoso , Grupo com Ancestrais do Continente Asiático/genética , Estudos de Casos e Controles , China/epidemiologia , Feminino , Frequência do Gene , Estudo de Associação Genômica Ampla , Humanos , Masculino , Pessoa de Meia-Idade , Razão de Chances , Medição de Risco/métodos , Neoplasias da Bexiga Urinária/etnologia , Neoplasias da Bexiga Urinária/genética
4.
Anticancer Res ; 40(5): 2537-2548, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32366398

RESUMO

BACKGROUND/AIM: Radiotherapy-induced autophagy affects radiation-sensitivity and radiotherapy efficacy. Histone modifications also occur during radiotherapy. This study assessed radiotherapy effects on histone modification and autophagy in non-small cell lung cancer (NSCLC) cells. MATERIALS AND METHODS: NSCLC cells were subjected to γ-irradiation. Autophagy was detected using western blotting and acridine orange staining. Radiation effect on cell growth was evaluated by clonogenic assay. Histone modifications were assessed by western blotting. Next generation sequencings (NGSs) were conducted to identify histone modification target genes. RESULTS: Radio-protective autophagy and histone H4 lysine 20 trimethylation (H4K20me3) were up-regulated after irradiation. By NGSs, genes that are differentially expressed upon irradiation were identified, including the candidate H4K20me3 target gene GABARAPL1. Furthermore, we showed that GABARAPL1 is essential for the radiation-induced autophagy. CONCLUSION: Our findings revealed the regulatory axis of radiation-induced H4K20me3-GABARAPL1 in radio-protective autophagy. Modulation of this axis may be a new strategy to enhance radiotherapy efficacy in NSCLC.


Assuntos
Autofagia/efeitos da radiação , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Histonas/metabolismo , Neoplasias Pulmonares/metabolismo , Lisina/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Biomarcadores Tumorais , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Carcinoma Pulmonar de Células não Pequenas/radioterapia , Linhagem Celular Tumoral , Epigênese Genética , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/radioterapia , Metilação , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , RNA Interferente Pequeno/genética
5.
DNA Cell Biol ; 39(6): 949-957, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32397755

RESUMO

Primary angle closure glaucoma (PACG) is a major cause of blindness worldwide. This study proposed to screen candidate PACG-associated variants in Chinese Han people. Whole exome sequencing was applied to five confirmed PACG patients and two primary angle closure suspect individuals within a PACG-enriched Chinese Han family. A series of bioinformatics analyses were implemented to obtain high-risk single nucleotide variant (SNV) loci for PACG, which were subsequently used for linkage analysis for identifying linkage genome regions. In addition, MassARRAY SNV genotyping was applied to high-risk PACG loci as well as those within linkage regions in another independent cohort including 251 PACG and 251 normal samples to further screen high-confidence SNVs. A total of 27 loci in 19 genes remained after linkage analysis. The 19 genes were significantly enriched in biological processes tightly related to PACG, including retinol metabolism and salmonella infection. Two nonsynonymous SNV loci, rs897804 in exon15 of HOOK2 and rs3745193 in exon7 of GTPBP3, were recognized with higher variant frequency in PACG samples than that in control samples after association analysis of MassARRAY SNV genotyping data. This study sheds new light on the understanding of PACG incidence among Chinese Han people.


Assuntos
Proteínas de Ligação ao GTP/genética , Predisposição Genética para Doença/genética , Glaucoma de Ângulo Fechado/genética , Proteínas Associadas aos Microtúbulos/genética , Polimorfismo de Nucleotídeo Único , Feminino , Loci Gênicos/genética , Humanos , Masculino , Linhagem , Sequenciamento Completo do Exoma
6.
PLoS One ; 15(5): e0232338, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32421702

RESUMO

End-binding proteins (EBs) associate with the growing microtubule plus ends to regulate microtubule dynamics as well as the interaction with intracellular structures. EB3 contributes to pathological vascular leakage through interacting with the inositol 1,4,5-trisphosphate receptor 3 (IP3R3), a calcium channel located at the endoplasmic reticulum membrane. The C-terminal domain of EB3 (residues 200-281) is functionally important for this interaction because it contains the effector binding sites, a prerequisite for EB3 activity and specificity. Structural data for this domain is limited. Here, we report the backbone chemical shift assignments for the human EB3 C-terminal domain and computationally explore its EB3 conformations. Backbone assignments, along with computational models, will allow future investigation of EB3 structural dynamics, interactions with effectors, and will facilitate the development of novel EB3 inhibitors.


Assuntos
Proteínas Associadas aos Microtúbulos/química , Ressonância Magnética Nuclear Biomolecular , Humanos , Modelos Moleculares , Domínios Proteicos , Estrutura Secundária de Proteína
7.
Parasitol Res ; 119(5): 1641-1652, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32285266

RESUMO

Clonorchis sinensis (C. sinensis) can induce a food-borne parasitic disease (clonorchiasis). Numerous studies have analyzed functional proteins, immunologic factors, pro-inflammatory cytokines, and cell signaling transduction that promote the development of clonorchiasis. In a previous study, it was shown that C. sinensis adult-derived total protein (CsTP) might be involved in the pathogenesis and development of liver fibrosis via bringing about Th2 immune response. In the present study, further investigation of CsTP on cellular function and inflammatory effect in vitro and in vivo has been elicited. CsTP induced inflammation and autophagy as evidenced by upregulation of TNF-α, IFN-γ, and autophagic markers LC3B and P62. Exposed to CsTP upregulated the antiapoptotic gene Bcl-2 expression, diminished the apoptosis induced by H2O2, but promoted the proliferation and migration of LX-2 cells in proper concentration range. Additionally, the protein levels of p-AKT and p-mTOR were repressed in response to CsTP, suggesting a correlation of blocking the activation of mTOR/AKT signaling pathway. These results revealed that CsTP might exacerbate hepatic pathological changes by regulating cell proliferation, apoptosis, autophagy, and inflammation in the liver and LX-2 cells. Some effects might be partially involved in the mTOR and AKT pathways.


Assuntos
Apoptose/fisiologia , Clonorquíase/patologia , Clonorchis sinensis/patogenicidade , Cirrose Hepática/patologia , Proteínas de Protozoários/metabolismo , Animais , Movimento Celular/fisiologia , Proliferação de Células/fisiologia , Clonorquíase/parasitologia , Clonorchis sinensis/genética , Citocinas/metabolismo , Doenças Transmitidas por Alimentos/parasitologia , Humanos , Peróxido de Hidrogênio/metabolismo , Inflamação/patologia , Interferon-alfa/metabolismo , Cirrose Hepática/parasitologia , Camundongos , Proteínas Associadas aos Microtúbulos/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/biossíntese , Proteínas de Ligação a RNA/metabolismo , Transdução de Sinais , Serina-Treonina Quinases TOR/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Regulação para Cima
8.
Nature ; 580(7801): 106-112, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32238932

RESUMO

Radial glial progenitor cells (RGPs) are the major neural progenitor cells that generate neurons and glia in the developing mammalian cerebral cortex1-4. In RGPs, the centrosome is positioned away from the nucleus at the apical surface of the ventricular zone of the cerebral cortex5-8. However, the molecular basis and precise function of this distinctive subcellular organization of the centrosome are largely unknown. Here we show in mice that anchoring of the centrosome to the apical membrane controls the mechanical properties of cortical RGPs, and consequently their mitotic behaviour and the size and formation of the cortex. The mother centriole in RGPs develops distal appendages that anchor it to the apical membrane. Selective removal of centrosomal protein 83 (CEP83) eliminates these distal appendages and disrupts the anchorage of the centrosome to the apical membrane, resulting in the disorganization of microtubules and stretching and stiffening of the apical membrane. The elimination of CEP83 also activates the mechanically sensitive yes-associated protein (YAP) and promotes the excessive proliferation of RGPs, together with a subsequent overproduction of intermediate progenitor cells, which leads to the formation of an enlarged cortex with abnormal folding. Simultaneous elimination of YAP suppresses the cortical enlargement and folding that is induced by the removal of CEP83. Together, these results indicate a previously unknown role of the centrosome in regulating the mechanical features of neural progenitor cells and the size and configuration of the mammalian cerebral cortex.


Assuntos
Centrossomo/metabolismo , Córtex Cerebral/citologia , Córtex Cerebral/embriologia , Células Ependimogliais/citologia , Células-Tronco Neurais/citologia , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Proteínas de Ciclo Celular/metabolismo , Membrana Celular/metabolismo , Membrana Celular/patologia , Proliferação de Células , Centríolos/metabolismo , Córtex Cerebral/patologia , Feminino , Masculino , Camundongos , Proteínas Associadas aos Microtúbulos/deficiência , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Microtúbulos/metabolismo , Microtúbulos/patologia , Neurogênese
9.
PLoS Pathog ; 16(4): e1008360, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32330185

RESUMO

Intestinal epithelial cells (IECs) are at the forefront of host-pathogen interactions, coordinating a cascade of immune responses to protect against pathogens. Here we show that IEC-intrinsic vitamin A signaling restricts pathogen invasion early in the infection and subsequently activates immune cells to promote pathogen clearance. Mice blocked for retinoic acid receptor (RAR) signaling selectively in IECs (stopΔIEC) showed higher Salmonella burden in colonic tissues early in the infection that associated with higher luminal and systemic loads of the pathogen at later stages. Higher pathogen burden in stopΔIEC mice correlated with attenuated mucosal interferon gamma (IFNγ) production by underlying immune cells. We found that, at homeostasis, the intestinal epithelium of stopΔIEC mice produced significantly lower amounts of interleukin 18 (IL-18), a potent inducer of IFNγ. Regulation of IL-18 by vitamin A was also observed in a dietary model of vitamin A supplementation. IL-18 reconstitution in stopΔIEC mice restored resistance to Salmonella by promoting epithelial cell shedding to eliminate infected cells and limit pathogen invasion early in infection. Further, IL-18 augmented IFNγ production by underlying immune cells to restrict pathogen burden and systemic spread. Our work uncovers a critical role for vitamin A in coordinating a biphasic immune response to Salmonella infection by regulating IL-18 production by IECs.


Assuntos
Microbioma Gastrointestinal , Interleucina-18/metabolismo , Mucosa Intestinal/imunologia , Proteínas Associadas aos Microtúbulos/fisiologia , Infecções por Salmonella/prevenção & controle , Salmonella typhimurium/imunologia , Vitamina A/metabolismo , Animais , Interações Hospedeiro-Patógeno , Interferon gama/metabolismo , Mucosa Intestinal/microbiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores do Ácido Retinoico/metabolismo , Infecções por Salmonella/imunologia , Infecções por Salmonella/microbiologia , Infecções por Salmonella/patologia , Transdução de Sinais
10.
Nat Cell Biol ; 22(5): 570-578, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32341547

RESUMO

Lissencephaly-1 (Lis1) is a key cofactor for dynein-mediated intracellular transport towards the minus-ends of microtubules. It remains unclear whether Lis1 serves as an inhibitor or an activator of mammalian dynein motility. Here we use single-molecule imaging and optical trapping to show that Lis1 does not directly alter the stepping and force production of individual dynein motors assembled with dynactin and a cargo adaptor. Instead, Lis1 promotes the formation of an active complex with dynactin. Lis1 also favours the recruitment of two dyneins to dynactin, resulting in increased velocity, higher force production and more effective competition against kinesin in a tug-of-war. Lis1 dissociates from motile complexes, indicating that its primary role is to orchestrate the assembly of the transport machinery. We propose that Lis1 binding releases dynein from its autoinhibited state, which provides a mechanistic explanation for why Lis1 is required for efficient transport of many dynein-associated cargos in cells.


Assuntos
Complexo Dinactina/metabolismo , Dineínas/metabolismo , Proteínas Associadas aos Microtúbulos/metabolismo , Animais , Linhagem Celular , Humanos , Cinesina/metabolismo , Microtúbulos/metabolismo , Ligação Proteica/fisiologia , Transporte Proteico/fisiologia , Células Sf9 , Suínos
11.
Nat Cell Biol ; 22(5): 559-569, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32341548

RESUMO

Dynein is a microtubule motor that transports many different cargos in various cell types and contexts. How dynein is regulated to perform these activities with spatial and temporal precision remains unclear. Human dynein is regulated by autoinhibition, whereby intermolecular contacts limit motor activity. Whether this mechanism is conserved throughout evolution, whether it can be affected by extrinsic factors, and its role in regulating dynein function remain unclear. Here, we use a combination of negative stain electron microscopy, single-molecule assays, genetic, and cell biological techniques to show that autoinhibition is conserved in budding yeast, and plays a key role in coordinating in vivo dynein function. Moreover, we find that the Lissencephaly-related protein, LIS1 (Pac1 in yeast), plays an important role in regulating dynein autoinhibition. Our studies demonstrate that, rather than inhibiting dynein motility, Pac1/LIS1 promotes dynein activity by stabilizing the uninhibited conformation, which ensures appropriate dynein localization and activity in cells.


Assuntos
1-Alquil-2-acetilglicerofosfocolina Esterase/metabolismo , Dineínas/metabolismo , Endorribonucleases/metabolismo , Proteínas Associadas aos Microtúbulos/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , Microtúbulos/metabolismo
12.
Nat Cell Biol ; 22(5): 518-525, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32341549

RESUMO

Cytoplasmic dynein-1 is a molecular motor that drives nearly all minus-end-directed microtubule-based transport in human cells, performing functions that range from retrograde axonal transport to mitotic spindle assembly1,2. Activated dynein complexes consist of one or two dynein dimers, the dynactin complex and an 'activating adaptor', and they show faster velocity when two dynein dimers are present3-6. Little is known about the assembly process of this massive ~4 MDa complex. Here, using purified recombinant human proteins, we uncover a role for the dynein-binding protein LIS1 in promoting the formation of activated dynein-dynactin complexes that contain two dynein dimers. Complexes activated by proteins representing three families of activating adaptors-BicD2, Hook3 and Ninl-all show enhanced motile properties in the presence of LIS1. Activated dynein complexes do not require sustained LIS1 binding for fast velocity. Using cryo-electron microscopy, we show that human LIS1 binds to dynein at two sites on the motor domain of dynein. Our research suggests that LIS1 binding at these sites functions in multiple stages of assembling the motile dynein-dynactin-activating adaptor complex.


Assuntos
1-Alquil-2-acetilglicerofosfocolina Esterase/metabolismo , Dineínas do Citoplasma/metabolismo , Complexo Dinactina/metabolismo , Proteínas Associadas aos Microtúbulos/metabolismo , Animais , Proteínas de Transporte/metabolismo , Células HEK293 , Humanos , Camundongos , Microtúbulos/metabolismo , Ligação Proteica/fisiologia , Proteínas Recombinantes/metabolismo
13.
PLoS One ; 15(4): e0224713, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32315296

RESUMO

Atrial metabolic disturbance contributes to the onset and development of atrial fibrillation (AF). Autophagy plays a role in maintaining the cellular energy balance. We examined whether atrial gene expressions related to fatty acid metabolism and autophagy are altered in chronic AF and whether they are related to each other. Right atrial tissue was obtained during heart surgery from 51 patients with sinus rhythm (SR, n = 38) or chronic AF (n = 13). Preoperative fasting serum free-fatty-acid levels were significantly higher in the AF patients. The atrial gene expression of fatty acid binding protein 3 (FABP3), which is involved in the cells' fatty acid uptake and intracellular fatty acid transport, was significantly increased in AF patients compared to SR patients; in the SR patients it was positively correlated with the right atrial diameter and intra-atrial electromechanical delay (EMD), parameters of structural and electrical atrial remodeling that were evaluated by an echocardiography. In contrast, the two groups' atrial contents of diacylglycerol (DAG), a toxic fatty acid metabolite, were comparable. Importantly, the atrial gene expression of microtubule-associated protein light chain 3 (LC3) was significantly increased in AF patients, and autophagy-related genes including LC3 were positively correlated with the atrial expression of FABP3. In conclusion, in chronic AF patients, the atrial expression of FABP3 was upregulated in association with autophagy-related genes without altered atrial DAG content. Our findings may support the hypothesis that dysregulated cardiac fatty acid metabolism contributes to the progression of AF and induction of autophagy has a cardioprotective effect against cardiac lipotoxicity in chronic AF.


Assuntos
Fibrilação Atrial/genética , Autofagia , Ácidos Graxos/metabolismo , Idoso , Fibrilação Atrial/metabolismo , Diglicerídeos/metabolismo , Proteína 3 Ligante de Ácido Graxo/genética , Proteína 3 Ligante de Ácido Graxo/metabolismo , Feminino , Átrios do Coração/metabolismo , Humanos , Masculino , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Pessoa de Meia-Idade , Regulação para Cima
15.
Life Sci ; 253: 117700, 2020 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-32335164

RESUMO

AIMS: Although previous studies elaborated that selective autophagy was involved in quality control of some organelles, including nucleus, mitochondria, the endoplasmic reticulum and peroxisomes, it remained unclear whether the selective autophagy of the Golgi apparatus (Golgiphagy) existed or not. MAIN METHODS: In this study, H9c2 cells, HUVECs, HA-VSMCs and HEK293T cells were treated with autophagy inducers, Golgi stress inducers and cardiomyocytes hypertrophy stimulators. The Golgiphagy was evaluated by analysing the co-localization of Golgi markers and LC3B. Furthermore, the transmission electron microscope was used to observe the occurrence of Golgiphagy. The co-immunoprecipitation assay was used to evaluate the interaction of GOLPH3 and LC3B. KEY FINDINGS: Results showed that starvation promoted the co-localization of both GM130-positive and TGN46-positive Golgi fragments with LC3B-positive autophagosomes in H9c2 cells, HUVECs, HA-VSMCs and HEK293T cells. Transmission electron microscopy images showed that Golgi apparatus was sequestered into the autophagosomes in the starvation group. Moreover, Golgi stress inducers also facilitated the co-localization of Golgi markers and LC3B in H9c2 cells, HUVECs, HA-VSMCs and HEK293T cells. Furthermore, cardiomyocyte hypertrophy stimulators also triggered the appearance of Golgiphagy in H9c2 cells. Importantly, the co-immunoprecipitation assay indicated endogenous GOLPH3 interacted with LC3B in H9c2 cells, HUVECs, HA-VSMCs. However, knocking down GOLPH3 inhibited the Golgiphagy. SIGNIFICANCE: This study unveiled a new selective autophagy of the Golgi apparatus (Golgiphagy). In addition, GOLPH3 might act as a novel cargo receptor to regulate Golgiphagy. Maintaining homeostasis of the Golgi apparatus via GOLPH3-mediated autophagy was indispensable for cell survival.


Assuntos
Autofagia/fisiologia , Complexo de Golgi/metabolismo , Proteínas de Membrana/metabolismo , Proteínas Associadas aos Microtúbulos/metabolismo , Animais , Linhagem Celular , Sobrevivência Celular/fisiologia , Técnicas de Silenciamento de Genes , Células HEK293 , Células Endoteliais da Veia Umbilical Humana , Humanos , Imunoprecipitação , Proteínas de Membrana/genética , Microscopia Eletrônica de Transmissão , Miócitos Cardíacos/metabolismo , Ratos
16.
Proc Natl Acad Sci U S A ; 117(16): 8876-8883, 2020 04 21.
Artigo em Inglês | MEDLINE | ID: mdl-32245812

RESUMO

Microtubules (MTs) are essential components of the eukaryotic cytoskeleton that serve as "highways" for intracellular trafficking. In addition to the well-known active transport of cargo by motor proteins, many MT-binding proteins seem to adopt diffusional motility as a transportation mechanism. However, because of the limited spatial resolution of current experimental techniques, the detailed mechanism of protein diffusion has not been elucidated. In particular, the precise role of tubulin tails and tail modifications in the diffusion process is unclear. Here, using coarse-grained molecular dynamics simulations validated against atomistic simulations, we explore the molecular mechanism of protein diffusion along MTs. We found that electrostatic interactions play a central role in protein diffusion; the disordered tubulin tails enhance affinity but slow down diffusion, and diffusion occurs in discrete steps. While diffusion along wild-type MT is performed in steps of dimeric tubulin, the removal of the tails results in a step of monomeric tubulin. We found that the energy barrier for diffusion is larger when diffusion on MTs is mediated primarily by the MT tails rather than the MT body. In addition, globular proteins (EB1 and PRC1) diffuse more slowly than an intrinsically disordered protein (Tau) on MTs. Finally, we found that polyglutamylation and polyglycylation of tubulin tails lead to slower protein diffusion along MTs, although polyglycylation leads to faster diffusion across MT protofilaments. Taken together, our results explain experimentally observed data and shed light on the roles played by disordered tubulin tails and tail modifications in the molecular mechanism of protein diffusion along MTs.


Assuntos
Difusão Facilitada/fisiologia , Microtúbulos/metabolismo , Tubulina (Proteína)/metabolismo , Proteínas de Ciclo Celular/metabolismo , Humanos , Proteínas Associadas aos Microtúbulos/metabolismo , Simulação de Dinâmica Molecular , Processamento de Proteína Pós-Traducional/fisiologia , Eletricidade Estática , Proteínas tau/metabolismo
17.
Nat Commun ; 11(1): 1781, 2020 04 14.
Artigo em Inglês | MEDLINE | ID: mdl-32286294

RESUMO

Polycomb Group (PcG) proteins form memory of transient transcriptional repression that is necessary for development. In Drosophila, DNA elements termed Polycomb Response Elements (PREs) recruit PcG proteins. How PcG activities are targeted to PREs to maintain repressed states only in appropriate developmental contexts has been difficult to elucidate. PcG complexes modify chromatin, but also interact with both RNA and DNA, and RNA is implicated in PcG targeting and function. Here we show that R-loops form at many PREs in Drosophila embryos, and correlate with repressive states. In vitro, both PRC1 and PRC2 can recognize R-loops and open DNA bubbles. Unexpectedly, we find that PRC2 drives formation of RNA-DNA hybrids, the key component of R-loops, from RNA and dsDNA. Our results identify R-loop formation as a feature of Drosophila PREs that can be recognized by PcG complexes, and RNA-DNA strand exchange as a PRC2 activity that could contribute to R-loop formation.


Assuntos
DNA/metabolismo , Proteínas de Drosophila/metabolismo , Histona-Lisina N-Metiltransferase/metabolismo , RNA/metabolismo , Animais , Drosophila , Proteínas de Drosophila/genética , Embrião não Mamífero/metabolismo , Inativação Gênica/fisiologia , Histona-Lisina N-Metiltransferase/genética , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Ligação Proteica
18.
J Cancer Res Clin Oncol ; 146(5): 1103-1113, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32157437

RESUMO

BACKGROUND: Microtubule-associated protein 1 light chain 3 (LC3), an autophagic gene, has been reported as a vital marker for many diseases and cancers. However, the role of LC3 in hepatocellular carcinoma (HCC) was not still investigated. Therefore, we conducted a meta-analysis to examine the association of LC3 with its clinicopathological and prognostic in HCC. METHODS: We consulted the PubMed, Cochrane Library, Web of Science, EMBASE, China National Knowledge Infrastructure and Wan Fang databases for published studies on LC3 in HCC. Newcastle-Ottawa scale was used to screen the quality of the literature. The statistical analysis was calculated by STATA 14.2. RESULTS: Of the 1329 titles identified, 10 articles involving 949 patients in HCC were included in this meta-analysis. The results of our study show that increased LC3 expression is related to size of tumor, but not to gender, age, number of tumor, liver cirrhosis, HBsAg, TNM stage, alpha fetoprotein, vascular invasion and histological grade. Positive LC3 expression was associated with overall survival by pooled hazard ratio. CONCLUSIONS: This meta-analysis indicated that positive LC3 expression was related to size of tumor, and could predict prognosis in human hepatocellular carcinoma.


Assuntos
Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Proteínas Associadas aos Microtúbulos/biossíntese , Autofagia/fisiologia , Humanos , Estadiamento de Neoplasias , Prognóstico
19.
Nat Commun ; 11(1): 1535, 2020 03 24.
Artigo em Inglês | MEDLINE | ID: mdl-32210230

RESUMO

Neurons maintain axonal homeostasis via employing a unique organization of the microtubule (MT) cytoskeleton, which supports axonal morphology and provides tracks for intracellular transport. Abnormal MT-based trafficking hallmarks the pathology of neurodegenerative diseases, but the exact mechanism regulating MT dynamics in axons remains enigmatic. Here we report on a regulation of MT dynamics by AuTophaGy(ATG)-related proteins, which previously have been linked to the autophagy pathway. We find that ATG proteins required for LC3 lipid conjugation are dispensable for survival of excitatory neurons and instead regulate MT stability via controlling the abundance of the MT-binding protein CLASP2. This function of ATGs is independent of their role in autophagy and requires the active zone protein ELKS1. Our results highlight a non-canonical role of ATG proteins in neurons and suggest that pharmacological activation of autophagy may not only promote the degradation of cytoplasmic material, but also impair axonal integrity via altering MT stability.


Assuntos
Autofagia/fisiologia , Axônios/fisiologia , Metabolismo dos Lipídeos/fisiologia , Microtúbulos/metabolismo , Animais , Animais Recém-Nascidos , Proteína 5 Relacionada à Autofagia/genética , Proteína 5 Relacionada à Autofagia/metabolismo , Sobrevivência Celular , Células Cultivadas , Técnicas de Inativação de Genes , Camundongos , Proteínas Associadas aos Microtúbulos/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Cultura Primária de Células , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Proteínas rab de Ligação ao GTP/metabolismo
20.
Zhongguo Xiu Fu Chong Jian Wai Ke Za Zhi ; 34(3): 318-322, 2020 Mar 15.
Artigo em Chinês | MEDLINE | ID: mdl-32174076

RESUMO

Objective: To investigate the expression and correlation of hypoxia inducible factor 1α (HIF-1α) and autophagy related molecules (Beclin1 and LC3B) in rat nucleus pulposus cells under hypoxia in vitro. Methods: The nucleus pulposus cells were extracted from the nucleus pulposus of healthy adult Sprague Dawley rats and passaged. The 3rd generation cells were identified by HE staining and collagenase type Ⅱ immunofluorescence staining and randomly divided into 4 groups. The cells in group A were cultured for 8 hours under normal oxygen condition (37℃, 5%CO 2, 20%O 2); the cells in group B were cultured for 8 hours under hypoxia condition (37℃, 5%CO 2, 1%O 2); the cells in group C were transfected with HIF-1α-small interfering RNA and cultured for 8 hours under hypoxia condition; and the cells in group D were cultured with autophagy inhibitor 3-MA for 8 hours under hypoxia condition. Western blot and real-time fluorescence quantitative PCR (qRT-PCR) were used to detect the expressions of HIF-1α and autophagy related molecules (Beclin1 and LC3B) in all groups. Results: HE staining of the 3rd generation nucleus pulposus cells showed that the cytoplasm was light pink and the nucleus was blue black, and the collagenase type Ⅱ immunofluorescence staining was positive. Western blot and qRT-PCR results showed that the relative expressions of HIF-1α, Beclin1, and LC3B proteins and genes in group B were significantly higher than those in group A ( P<0.05); the relative expressions of HIF-1α, Beclin1, and LC3B proteins and genes in group C were significantly lower than those in group B ( P<0.05). There was no significant difference in the relative expression of HIF-1α protein and gene between groups B and D ( P>0.05); while the relative expressions of Beclin1 and LC3B proteins and genes in group D were significant lower than those in group B ( P<0.05). Conclusion: Hypoxia can induce the expressions of HIF-1α and autophagy related molecules (Beclin1 and LC3B) in rat nucleus pulposus cells, and HIF-1α in rat nucleus pulposus cells under hypoxia is related to the expression of autophagy related molecules, that is, down-regulation of HIF-1α can significantly reduce the expression of autophagy related molecules, while the down-regulation of autophagy levels under hypoxia has no or little effect on the expression of HIF-1α.


Assuntos
Autofagia , Proteína Beclina-1/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Proteínas Associadas aos Microtúbulos/metabolismo , Núcleo Pulposo/citologia , Animais , Hipóxia Celular , Células Cultivadas , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley
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