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1.
Results Probl Cell Differ ; 67: 27-48, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31435791

RESUMO

Formin homology proteins (formins) are a highly conserved family of cytoskeletal remodeling proteins that are involved in a diverse array of cellular functions. Formins are best known for their ability to regulate actin dynamics, but the same functional domains also govern stability and organization of microtubules. It is thought that this dual activity allows them to coordinate the activity of these two major cytoskeletal networks and thereby influence cellular architecture. Golgi ribbon assembly is dependent upon cooperative interactions between actin filaments and cytoplasmic microtubules originating both at the Golgi itself and from the centrosome. Similarly, centrosome assembly, centriole duplication, and centrosome positioning are also reliant on a dialogue between both cytoskeletal networks. As presented in this chapter, a growing body of evidence suggests that multiple formin proteins play essential roles in these central cellular processes.


Assuntos
Centríolos/metabolismo , Proteínas do Citoesqueleto/metabolismo , Citoesqueleto/metabolismo , Complexo de Golgi/metabolismo , Citoesqueleto de Actina/metabolismo , Actinas/metabolismo , Microtúbulos/metabolismo
2.
Results Probl Cell Differ ; 67: 81-93, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31435793

RESUMO

In vertebrate cells, the Golgi apparatus is located in close proximity to the centriole. The architecture of the Golgi/centriole complex depends on a multitude of factors, including the actin filament cytoskeleton. In turn, both the Golgi and centriole act as the actin nucleation centers. Actin organization and polymerization also depend on the small GTPase RhoA pathway. In this chapter, we summarize the most current knowledge on how the genetic, magnetic, or pharmacologic interference with RhoA pathway and actin cytoskeleton directly or indirectly affects architecture, structure, and function of the Golgi/centriole complex.


Assuntos
Actinas/metabolismo , Centríolos/metabolismo , Complexo de Golgi/metabolismo , Proteína rhoA de Ligação ao GTP/metabolismo , Citoesqueleto de Actina/metabolismo
3.
J Agric Food Chem ; 67(35): 9789-9795, 2019 Sep 04.
Artigo em Inglês | MEDLINE | ID: mdl-31373816

RESUMO

Pulmonary fibrosis is a chronic lung disease characterized by abnormal accumulation of the extracellular matrix (ECM). Chronic damage of the alveolar epithelium leads to a process called "epithelial-mesenchymal transition" (EMT) and increases synthesis and deposition of ECM proteins. Therefore, inhibition of EMT might be a promising therapeutic approach for the treatment of pulmonary fibrosis. ß-Sitosterol is one of the most abundant phytosterols in the plant kingdom and the major constituent in corn silk, which is derived from the stigma and style of maize (Zea mays). In this study, we elucidated that ß-sitosterol inhibited transforming growth factor-ß1 (TGF-ß1)-induced EMT and consequently had an antifibrotic effect. ß-Sitosterol (1-10 µg/mL) significantly downregulated the TGF-ß1-induced fibrotic proteins, such as collagen, fibronectin, and α-smooth muscle actin in human alveolar epithelial cells (p < 0.01). After 24 h, relative wound density (RWD) was increased in TGF-ß1 treated group (82.16 ± 5.70) compare to the control group (64.63 ± 2.21), but RWD was decreased in ß-sitosterol cotreated group (10 µg/mL: 71.54 ± 7.39; 20 µg/mL: 65.69 ± 6.42). In addition, the changes of the TGF-ß1-induced morphological shape and protein expression of EMT markers, N-cadherin, vimentin, and E-cadherin, were significantly blocked by ß-sitosterol treatment (p < 0.01). The effects of ß-sitosterol on EMT were found to be associated with the TGF-ß1/Snail pathway, which is regulated by Smad and non-Smad signaling pathways. Taken together, these findings suggest that ß-sitosterol can be used to attenuate pulmonary fibrosis through suppression of EMT by inhibiting the TGF-ß1/Snail pathway.


Assuntos
Células Epiteliais Alveolares/efeitos dos fármacos , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Extratos Vegetais/farmacologia , Alvéolos Pulmonares/efeitos dos fármacos , Fibrose Pulmonar/fisiopatologia , Sitosteroides/farmacologia , Zea mays/química , Actinas/genética , Actinas/metabolismo , Células Epiteliais Alveolares/citologia , Células Epiteliais Alveolares/metabolismo , Caderinas/genética , Caderinas/metabolismo , Linhagem Celular , Fibronectinas/genética , Fibronectinas/metabolismo , Humanos , Extratos Vegetais/química , Alvéolos Pulmonares/metabolismo , Alvéolos Pulmonares/fisiopatologia , Fibrose Pulmonar/tratamento farmacológico , Fibrose Pulmonar/genética , Fibrose Pulmonar/metabolismo , Transdução de Sinais/efeitos dos fármacos , Fator de Crescimento Transformador beta1/genética , Fator de Crescimento Transformador beta1/metabolismo
4.
Zhong Nan Da Xue Xue Bao Yi Xue Ban ; 44(5): 485-490, 2019 May 28.
Artigo em Chinês | MEDLINE | ID: mdl-31303610

RESUMO

OBJECTIVE: To explore the magnetic resonance imaging (MRI) characteristics of glioma with Brg/Brm-associated factor 53a (BAF53a) expression.
 Methods: A total of 121 patients with glioma was divided into a BAF53a high expression group (n=79) and a low expression group (n=42) according to the results of immunohistochemistry. Then the MRI characteristics, including lesion location, number, boundary, maximum diameter, peripheral edema, midline structure shift, homogeneity, cystic necrosis, hemorrhage, strengthening degree, ependymal strengthening, pia mater enhancement, deep white matter invasion and lesion across the midline (total 14 items), were analyzed.
 Results: The results showed that there were significance difference in lesion border, lesion edema, enhancement of the lesion, and deep white matter invasion between the 2 groups (all P<0.05).
 Conclusion: The MRI characteristics, such as lesion border, lesion edema degree, enhancement degree of the lesion and deep white matter invasion, might be associated with BAF53a expression in gliomas.


Assuntos
Actinas/metabolismo , Neoplasias Encefálicas , Proteínas Cromossômicas não Histona/metabolismo , Proteínas de Ligação a DNA/metabolismo , Glioma , Humanos , Imagem por Ressonância Magnética , Necrose
5.
Biomed Environ Sci ; 32(6): 419-426, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31262387

RESUMO

OBJECTIVE: Silicosis, caused by inhalation of silica dust, is the most serious occupational disease in China and the aim of present study was to explore the protective effect of Ang (1-7) on silicotic fibrosis and myofibroblast differentiation induced by Ang II. METHODS: HOPE-MED 8050 exposure control apparatus was used to establish the rat silicosis model. Pathological changes and collagen deposition of the lung tissue were examined by H.E. and VG staining, respectively. The localizations of ACE2 and α-smooth muscle actin (α-SMA) in the lung were detected by immunohistochemistry. Expression levels of collagen type I, α-SMA, ACE2, and Mas in the lung tissue and fibroblasts were examined by western blot. Levels of ACE2, Ang (1-7), and Ang II in serum were determined by ELISA. Co-localization of ACE2 and α-SMA in fibroblasts was detected by immunofluorescence. RESULTS: Ang (1-7) induced pathological changes and enhanced collagen deposition in vivo. Ang (1-7) decreased the expressions of collagen type I and α-SMA and increased the expressions of ACE2 and Mas in the silicotic rat lung tissue and fibroblasts stimulated by Ang II. Ang (1-7) increased the levels of ACE2 and Ang (1-7) and decreased the level of Ang II in silicotic rat serum. A779 enhanced the protective effect of Ang (1-7) in fibroblasts stimulated by Ang II. CONCLUSION: Ang (1-7) exerted protective effect on silicotic fibrosis and myofibroblast differentiation induced by Ang II by regulating ACE2-Ang (1-7)-Mas axis.


Assuntos
Angiotensina II/sangue , Angiotensina I/uso terapêutico , Pulmão/metabolismo , Miofibroblastos/efeitos dos fármacos , Fragmentos de Peptídeos/uso terapêutico , Silicose/prevenção & controle , Actinas/metabolismo , Angiotensina I/sangue , Angiotensina I/farmacologia , Animais , Animais Recém-Nascidos , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Colágeno Tipo I/metabolismo , Modelos Animais de Doenças , Pulmão/patologia , Fragmentos de Peptídeos/sangue , Fragmentos de Peptídeos/farmacologia , Peptidil Dipeptidase A/metabolismo , Ratos Wistar , Silicose/metabolismo , Silicose/patologia
6.
Life Sci ; 232: 116637, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31288014

RESUMO

Keloid is characterized by overactive fibroblasts. Forkhead box M1 (FOXM1) is transcription factor that plays important roles in the progression of fibrosis. However, the role of FOXM1 in keloid has not been elucidated. In the present study, we examined the expression levels of FOXM1 in clinical keloid tissue specimens and primary keloid fibroblasts (KFs). The results showed that FOXM1 levels were significantly increased in both keloid tissues and KFs. To further investigate the biological functions of FOXM1, FOXM1 was knocked down in KFs by transfection with small interfering RNA targeting FOXM1 (si-FOXM1). Knockdown of FOXM1 inhibited transforming growth factor-ß1 (TGF-ß1)-induced cell proliferation and migration of KFs. Besides, the increased expressions of collagen (coll I), connective tissue growth factor (CTGF), and α-smooth muscle actin (α-SMA) in TGF-ß1-induced KFs were suppressed by si-FOXM1 transfection. Furthermore, TGF-ß1-induced increase in p-Smad2 and p-Smad3 expressions was attenuated by FOXM1 knockdown. These data indicated that knockdown of FOXM1 inhibited TGF-ß1-induced KFs activation and extracellular matrix (ECM) accumulation, which was attributed to the inhibition of TGF-ß1/Smad pathway.


Assuntos
Proteína Forkhead Box M1/deficiência , Queloide/metabolismo , Actinas/metabolismo , Adolescente , Adulto , Movimento Celular/fisiologia , Proliferação de Células/fisiologia , Células Cultivadas , Colágeno/metabolismo , Colágeno Tipo I/metabolismo , Fator de Crescimento do Tecido Conjuntivo/metabolismo , Matriz Extracelular/genética , Matriz Extracelular/metabolismo , Feminino , Fibroblastos/metabolismo , Proteína Forkhead Box M1/genética , Proteína Forkhead Box M1/metabolismo , Técnicas de Silenciamento de Genes/métodos , Humanos , Queloide/genética , Masculino , Fosforilação , RNA Interferente Pequeno/genética , Transdução de Sinais , Proteína Smad2/antagonistas & inibidores , Proteína Smad2/metabolismo , Proteína Smad3/antagonistas & inibidores , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta1/antagonistas & inibidores , Fator de Crescimento Transformador beta1/metabolismo
7.
Vet Microbiol ; 235: 127-135, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31282370

RESUMO

Lawsonia intracellularis is an obligate intracellular Gram-negative bacterium that has been identified as the etiological agent of the contagious disease proliferative enteropathy (PE) in a wide range of animals, mainly pigs. The genome sequence of L. intracellularis indicates that this bacterium possess a type III secretion system (T3SS), which may assist the bacterium during cell invasion and host innate immune system evasion and could be a mechanism for inducing cellular proliferation. However, the effectors secreted by the T3SS (T3Es) of L. intracellularis have not been reported. T3Es often target conserved eukaryotic cellular processes, and yeast is an established and robust model system in which to reveal their function. By screening the growth inhibition of an ordered array of Saccharomyces cerevisiae strains expressing the hypothetical genes of L. intracellularis, LI1035 was identified as the first putative effector that inhibits yeast growth. The LI1035-induced growth inhibition was rescued in two of the 14 mitogen-activated protein kinase (MAPK) yeast haploid deletion strains, suggesting that LI1035 interacts with the components of the MAPK pathway in yeast. Phosphorylation assays confirmed that LI1035 inhibits MAPK signaling cascades in yeast and mammalian cells. Actin staining assays revealed that LI1035 regulates actin organization in yeast and mammalian cells. Taken together, these results indicate that LI1035 alters MAPK pathway activity and regulates actin organization in the host. These findings may contribute to the understanding the pathogenesis of L. intracellularis and support the use of yeast as a heterologous system for the functional analysis of pathogen-specific gene products in the laboratory.


Assuntos
Actinas/metabolismo , Proteínas de Bactérias/metabolismo , Lawsonia (Bactéria)/metabolismo , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Saccharomyces cerevisiae/crescimento & desenvolvimento , Transdução de Sinais , Animais , Proteínas de Bactérias/genética , Proliferação de Células , Interações entre Hospedeiro e Microrganismos , Lawsonia (Bactéria)/genética , Fosforilação , Saccharomyces cerevisiae/genética , Sorbitol/farmacologia , Suínos , Temperatura Ambiente
8.
Life Sci ; 231: 116674, 2019 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-31344427

RESUMO

Hypertrophic scar formation is a fibroproliferative disorder caused by abnormal wound healing. At present, there are limited treatment strategies for hypertrophic scars. In this study, we identified an endogenous peptide, LYENRL, through peptidomics screening that is downregulated in scar skin tissues. The peptide exhibited concentration dependent inhibitory effects on the proliferation, migration and extracellular matrix (ECM) production of scar fibroblasts. By eukaryotic transcriptome sequencing analysis, we noted that LYENRL downregulated gene sets in scar fibroblasts were associated with the transforming growth factor-ß (TGF-ß) signaling pathway. Further experiments revealed that LYENRL was able to inhibit the activation of TGF-ß1/Smad signaling and TGF-ß1-induced activation of scar fibroblasts at the source by blocking the binding of AP-1 to the corresponding region of the Tgfb1 promoter, which in turn inhibited gene expression of Tgfb1. Taken together, we concluded that the effects of LYENRL on scar fibroblasts make it a potential peptide drug for hypertrophic scar treatment.


Assuntos
Cicatriz Hipertrófica/metabolismo , Cicatriz Hipertrófica/patologia , Peptídeos/farmacologia , Actinas/metabolismo , Linhagem Celular , Células Cultivadas , Regulação para Baixo/efeitos dos fármacos , Matriz Extracelular/metabolismo , Feminino , Fibroblastos/efeitos dos fármacos , Fibroblastos/fisiologia , Humanos , Peptídeos/metabolismo , Cultura Primária de Células , Transdução de Sinais/efeitos dos fármacos , Pele/metabolismo , Proteínas Smad/metabolismo , Proteínas Smad/fisiologia , Proteína Smad3/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Fator de Crescimento Transformador beta1/fisiologia
9.
Biochemistry (Mosc) ; 84(4): 358-369, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31228927

RESUMO

Cytoplasmic actin structures are essential components of the eukaryotic cytoskeleton. According to the classic concepts, actin structures perform contractile and motor functions, ensuring the possibility of cell shape changes during cell spreading, polarization, and movement both in vitro and in vivo, from the early embryogenesis stages and throughout the life of a multicellular organism. Intracellular organization of actin structures, their biochemical composition, and dynamic properties play a key role in the realization of specific cellular and tissue functions and vary in different cell types. This paper is a review of recent studies on the organization and properties of actin structures in endotheliocytes, interaction of these structures with other cytoskeletal components and elements involved in cell adhesion, as well as their role in the functional activity of endothelial cells.


Assuntos
Citoesqueleto de Actina/metabolismo , Actinas/metabolismo , Citoesqueleto de Actina/química , Actinas/química , Actinas/genética , Caderinas/química , Caderinas/metabolismo , Citosol/metabolismo , Células Endoteliais/citologia , Células Endoteliais/metabolismo , Humanos , Proteínas Associadas aos Microtúbulos/metabolismo , Microtúbulos/química , Microtúbulos/metabolismo
10.
Pol J Microbiol ; 68(2): 217-224, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31250592

RESUMO

Campylobacter fetus is an important venereal pathogen of cattle that causes infertility and abortions. It is transmitted during mating, and it travels from the vagina to the uterus; therefore, an important cell type that interacts with C. fetus are endometrial epithelial cells. Several virulence factors have been identified in the genome of C. fetus, such as adhesins, secretion systems, and antiphagocytic layers, but their expression is unknown. The ability of C. fetus to invade human epithelial cells has been demonstrated, but the ability of this microorganism to infect bovine endometrial epithelial cells has not been demonstrated. Bovine endometrial epithelial cells were isolated and challenged with C. fetus. The presence of C. fetus inside the endometrial epithelial cells was confirmed by the confocal immunofluorescence. C. fetus was not internalized when actin polymerization was disturbed, suggesting cytoskeleton participation in an internalization mechanism. To evaluate the intracellular survival of C. fetus, a gentamicin protection assay was performed. Although C. fetus was able to invade epithelial cells, the results showed that it did not have the capacity to survive in the intracellular environment. This study reports for the first time, the ability of C. fetus to invade bovine endometrial epithelial cells, and actin participation in this phenomenon.Campylobacter fetus is an important venereal pathogen of cattle that causes infertility and abortions. It is transmitted during mating, and it travels from the vagina to the uterus; therefore, an important cell type that interacts with C. fetus are endometrial epithelial cells. Several virulence factors have been identified in the genome of C. fetus, such as adhesins, secretion systems, and antiphagocytic layers, but their expression is unknown. The ability of C. fetus to invade human epithelial cells has been demonstrated, but the ability of this microorganism to infect bovine endometrial epithelial cells has not been demonstrated. Bovine endometrial epithelial cells were isolated and challenged with C. fetus. The presence of C. fetus inside the endometrial epithelial cells was confirmed by the confocal immunofluorescence. C. fetus was not internalized when actin polymerization was disturbed, suggesting cytoskeleton participation in an internalization mechanism. To evaluate the intracellular survival of C. fetus, a gentamicin protection assay was performed. Although C. fetus was able to invade epithelial cells, the results showed that it did not have the capacity to survive in the intracellular environment. This study reports for the first time, the ability of C. fetus to invade bovine endometrial epithelial cells, and actin participation in this phenomenon.


Assuntos
Infecções por Campylobacter/microbiologia , Campylobacter fetus/fisiologia , Endocitose , Células Epiteliais/microbiologia , Actinas/metabolismo , Animais , Antibacterianos/farmacologia , Bovinos , Doenças dos Bovinos , Células Cultivadas , Gentamicinas/farmacologia , Viabilidade Microbiana/efeitos dos fármacos , Microscopia Confocal , Microscopia de Fluorescência , Modelos Biológicos
11.
Nat Commun ; 10(1): 2685, 2019 06 18.
Artigo em Inglês | MEDLINE | ID: mdl-31213605

RESUMO

Hypertrophic cardiomyopathy (HCM) affects 1 in 500 people and leads to hyper-contractility of the heart. Nearly 40 percent of HCM-causing mutations are found in human ß-cardiac myosin. Previous studies looking at the effect of HCM mutations on the force, velocity and ATPase activity of the catalytic domain of human ß-cardiac myosin have not shown clear trends leading to hypercontractility at the molecular scale. Here we present functional data showing that four separate HCM mutations located at the myosin head-tail (R249Q, H251N) and head-head (D382Y, R719W) interfaces of a folded-back sequestered state referred to as the interacting heads motif (IHM) lead to a significant increase in the number of heads functionally accessible for interaction with actin. These results provide evidence that HCM mutations can modulate myosin activity by disrupting intramolecular interactions within the proposed sequestered state, which could lead to hypercontractility at the molecular level.


Assuntos
Miosinas Cardíacas/metabolismo , Cardiomiopatia Hipertrófica/genética , Cardiomiopatia Hipertrófica/fisiopatologia , Contração Miocárdica/genética , Cadeias Pesadas de Miosina/metabolismo , Actinas/metabolismo , Animais , Miosinas Cardíacas/genética , Linhagem Celular , Movimento Celular/genética , Coração/fisiopatologia , Humanos , Camundongos , Mutação , Mioblastos , Cadeias Pesadas de Miosina/genética , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo
12.
Nat Commun ; 10(1): 2856, 2019 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-31253774

RESUMO

Microfilaments (actin) and microtubules represent the extremes in eukaryotic cytoskeleton cross-sectional dimensions, raising the question of whether filament architectures are limited by protein fold. Here, we report the cryoelectron microscopy structure of a complex filament formed from 15 protofilaments of an actin-like protein. This actin-like ParM is encoded on the large pCBH Clostridium botulinum plasmid. In cross-section, the ~26 nm diameter filament comprises a central helical protofilament surrounded by intermediate and outer layers of six and eight twisted protofilaments, respectively. Alternating polarity of the layers allows for similar lateral contacts between each layer. This filament design is stiffer than the actin filament, and has likely been selected for during evolution to move large cargos. The comparable sizes of microtubule and pCBH ParM filaments indicate that larger filament architectures are not limited by the protomer fold. Instead, function appears to have been the evolutionary driving force to produce broad, complex filaments.


Assuntos
Actinas/metabolismo , Proteínas de Bactérias/metabolismo , Clostridium botulinum/metabolismo , Citoesqueleto/fisiologia , Citoesqueleto de Actina , Actinas/genética , Proteínas de Bactérias/genética , Microscopia Crioeletrônica , Regulação Bacteriana da Expressão Gênica/fisiologia , Modelos Moleculares , Conformação Proteica
13.
Histochem Cell Biol ; 152(2): 133-143, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31154480

RESUMO

Actin fulfills important cytoplasmic but also nuclear functions in eukaryotic cells. In the nucleus, actin modulates gene expression and chromatin remodeling. Monomeric (G-actin) and polymerized actin (F-actin) have been analyzed by fluorescence microscopy in the nucleus; however, the resolution at the ultrastructural level has not been investigated in great detail. We provide a first documentation of nuclear actin in mouse fibroblasts by electron microscopy (EM). For this, we employed correlative light and electron microscopy on the same section using actin-directed nanobodies recognizing endogenous monomeric and polymeric actin proteins (so-called nuclear Actin-chromobody-GFP; nAC-GFP). Indeed, using this strategy, we could identify actin proteins present in the nucleus. Here, immunogold-labeled actin proteins were spread throughout the entire nucleoplasm. Of note, nuclear actin was complementarily localized to DAPI-positive areas, the latter marking preferentially transcriptionally inactive heterochromatin. Since actin aggregates in rod structures upon cell stress including neurodegeneration, we analyzed nuclear actin at the ultrastructural level after DMSO or UV-mediated cell damage. In those cells the ratio between cytoplasmic and nuclear gold-labeled actin proteins was altered compared to untreated control cells. In summary, this EM analysis (i) confirmed the presence of endogenous nuclear actin at ultrastructural resolution, (ii) revealed the actin abundance in less chromatin-dense regions potentially reflecting more transcriptionally active euchromatin rather than transcriptionally inactive heterochromatin and (iii) showed an altered abundance of actin-associated gold particles upon cell stress.


Assuntos
Actinas/análise , Núcleo Celular/química , Microscopia Eletrônica/métodos , Microscopia de Fluorescência/métodos , Actinas/metabolismo , Animais , Núcleo Celular/metabolismo , Células Cultivadas , Fibroblastos/química , Fibroblastos/citologia , Camundongos , Células NIH 3T3 , Tamanho da Partícula , Conformação Proteica
14.
Life Sci ; 232: 116591, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31228513

RESUMO

AIMS: Calreticulin (CRT) is a multifunctional protein that protects endothelial cells by alleviating actin cytoskeleton injury, but the underlying mechanism remains unclear. CRT was recently identified as a novel acyltransferase; acetylation at the N-terminus of actin monomers strengthens actin polymerization. This study was undertaken to determine whether CRT protects human microvascular endothelial cells (HMECs) against microwave radiation through actin acetylation. MATERIALS AND METHODS: We prepared a eukaryotic-derived recombinant CRT and incubated the HMECs with it prior to microwave exposure. We then assessed cell injury and endothelial function, detected actin polymerization and acetylation after HMECs exposure to S-band high-power microwaves. Coimmunoprecipitation, pull-down, and ex vitro acetylation reaction were performed to determine whether actin is a novel substrate of CRT acyltransferase. Finally, we employed the mutant experiments to demonstrate the acetylation sites contributing to CRT acetyltransferase activity. KEY FINDINGS: Microwave radiation induced severe cell injury and endothelial contact dysfunction, reduced the polymerization of actin filaments, and destroyed the actin arrangement, ultimately reducing acetylated actin expression. CRT treatment upregulated actin acetylation levels, promoted polymerization, and facilitated thicker and longer F-actin stress fibre formation. Pre-incubation with CRT rescued microwave-induced cell injury, decreased actin acetylation, and rendered the actin cytoskeleton radiation-retardant. The level of acetyl-actin was positively correlated with actin polymerization. Actin was identified as a novel substrate of CRT, being acetylated mainly through the CRT P-domain at lys-206 and -207. SIGNIFICANCE: This work provides a better understanding of the underlying mechanism of CRT-induced cytoprotection, and suggests a novel therapeutic target for microwave radiation-related diseases with endothelial dysfunction.


Assuntos
Actinas/metabolismo , Calreticulina/farmacologia , Microvasos/efeitos dos fármacos , Acetilação/efeitos dos fármacos , Citoesqueleto de Actina/metabolismo , Linhagem Celular , Citoproteção , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Humanos , Microvasos/efeitos da radiação , Micro-Ondas , Substâncias Protetoras , Processamento de Proteína Pós-Traducional , Transdução de Sinais
15.
Nat Commun ; 10(1): 2356, 2019 05 29.
Artigo em Inglês | MEDLINE | ID: mdl-31142743

RESUMO

Centrosomes control cell motility, polarity and migration that is thought to be mediated by their microtubule-organizing capacity. Here we demonstrate that WNT signalling drives a distinct form of non-directional cell motility that requires a key centrosome module, but not microtubules or centrosomes. Upon exosome mobilization of PCP-proteins, we show that DVL2 orchestrates recruitment of a CEP192-PLK4/AURKB complex to the cell cortex where PLK4/AURKB act redundantly to drive protrusive activity and cell motility. This is mediated by coordination of formin-dependent actin remodelling through displacement of cortically localized DAAM1 for DAAM2. Furthermore, abnormal expression of PLK4, AURKB and DAAM1 is associated with poor outcomes in breast and bladder cancers. Thus, a centrosomal module plays an atypical function in WNT signalling and actin nucleation that is critical for cancer cell motility and is associated with more aggressive cancers. These studies have broad implications in how contextual signalling controls distinct modes of cell migration.


Assuntos
Aurora Quinase B/metabolismo , Movimento Celular , Centrossomo/metabolismo , Proteínas Cromossômicas não Histona/metabolismo , Proteínas Desgrenhadas/metabolismo , Neoplasias/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Via de Sinalização Wnt , Actinas/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Neoplasias da Mama/metabolismo , Carcinoma de Células de Transição/metabolismo , Linhagem Celular Tumoral , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas dos Microfilamentos/metabolismo , Prognóstico , Mapas de Interação de Proteínas , Reação em Cadeia da Polimerase em Tempo Real , Neoplasias da Bexiga Urinária/metabolismo
16.
Phys Rev E ; 99(4-1): 042417, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31108612

RESUMO

Cellular checkerboard patterns are observed at many stages of embryonic development. We study an analytically tractable model for lateral inhibition and show that the steady states are analogous to optical phonons at the Γ point, which have the wave number k=0. We study the cases of cells arranged in linear and hexagonal lattices. To determine how the final pattern is selected it is necessary to take into account the granularity of the pattern and, analogously to solid-state physics, to redefine the basis and lattice sites in terms of a periodic crystal. The sites and basis are determined by looking at the symmetries of inhibitory interactions between cells. The redefined basis for cells placed in a linear lattice is composed by two cells which are embedded in another linear lattice, while for cells placed in a hexagonal lattice the redefined basis consists of three cells embedded in another hexagonal lattice. The pattern in hexagonal lattices can be driven into three different states: two of those states are periodic checkerboards and a third in which both periodic states coexist. These observations provides new predictions for experiments.


Assuntos
Desenvolvimento Embrionário , Modelos Biológicos , Actinas/metabolismo , Fenômenos Ópticos
17.
J Med Microbiol ; 68(6): 940-951, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31107199

RESUMO

PURPOSE: This study aimed to characterize 82 atypical enteropathogenic Escherichia coli (aEPEC) isolates, obtained from patients with diarrhea in Brazil, regarding their adherence patterns on HeLa cells and attaching and effacing (AE) lesion pathways. METHODOLOGY: The adherence and fluorescence-actin staining (FAS) assays were performed using HeLa cells. AE lesion pathways were determined through the detection of tyrosine residue 474 (Y474) phosphorylation in the Tir protein, after its translocation to host cells, and by PCR assays for tir genotyping and detection of Tir-cytoskeleton coupling protein (tccP) genes. RESULTS: Regarding the adherence pattern, determined in the presence of d-mannose, 12 isolates (14.6 %) showed the localized adherence (LA)-like pattern, 3 (3.7  %) the aggregative adherence pattern and 4 (4.9  %) a hybrid LA/diffuse adherence pattern. In addition, 36 (43.9  %) isolates displayed an undefined adherence, and 26 (31.7  %) were non-adherent (NA), while one (1.2 %) caused cell detachment. Among the 26 NA aEPEC isolates, 11 showed a type 1 pilus-dependent adherence in assays performed without d-mannose, while 15 remained NA. Forty-eight (58.5 %) aEPEC were able to trigger F-actin accumulation underneath adherent bacteria (FAS-positive), which is an important feature of AE lesions. The majority (58.3 %) of these used the Tir-Nck pathway, while 39.6  % may use both Tir-Nck and Tir-TccP pathways to induce AE lesions. CONCLUSION: Our results reveal the diversity of strategies used by aEPEC isolates to interact with and damage epithelial host cells, thereby causing diarrheal diseases.


Assuntos
Aderência Bacteriana , Escherichia coli Enteropatogênica/fisiologia , Infecções por Escherichia coli/microbiologia , Interações Hospedeiro-Patógeno , Actinas/metabolismo , Diarreia/microbiologia , Escherichia coli Enteropatogênica/genética , Escherichia coli Enteropatogênica/isolamento & purificação , Células Epiteliais/microbiologia , Proteínas de Escherichia coli/metabolismo , Fímbrias Bacterianas/metabolismo , Genótipo , Células HeLa , Humanos , Fenótipo , Fosforilação , Receptores de Superfície Celular/metabolismo
18.
Yonsei Med J ; 60(6): 509-516, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31124333

RESUMO

PURPOSE: This study was conducted to verify the induction and mechanism of selective apoptosis in G361 melanoma cells using anti-HER2 antibody-conjugated gold nanoparticles (GNP-HER2). MATERIALS AND METHODS: Following GNP-HER2 treatment of G361 cells, cell cycle arrest and apoptosis were measured by WST-1 assay, Hemacolor staining, Hoechst staining, immunofluorescence staining, fluorescence-activated cell sorting analysis, and Western blotting. RESULTS: G361 cells treated with GNP-HER2 showed condensation of nuclei, which is an apoptotic phenomenon, and translocation of apoptosis-inducing factor and cytochrome c from mitochondria into the nucleus and cytoplasm, respectively. Increases in BAX in cells undergoing apoptosis, activation of caspase-3 and -9, and fragmentation of poly (ADP-ribose) polymerase and DNA fragmentation factor 45 (inhibitor of caspase-activated DNase) were observed upon GNP-HER2 treatment. Following GNP-HER2 treatment, an increase of cells in sub-G1 phase, which is a signal of cell apoptosis, was observed. This resulted in the down-regulation of cyclin A, cyclin D1, cyclin E, cdk2, cdk4, and cdc2 and the up-regulation of p21. Thus, GNP-HER2 treatment was confirmed to induce the cessation of cell cycle progression. Also, decreases in phospho-focal adhesion kinase and phospho-human epidermal growth factor receptor, which activate cellular focal adhesion, and decreases in phospho-paxillin, which stimulates the disassembly of filamentous actin, were observed. Reduced cell adhesion and disassembly of the intracellular structure indicated cell deactivation. CONCLUSION: GNP-HER2 can selectively kill G361 melanoma cells without affecting normal cells. The mechanism of G361 cell death upon treatment with GNP-HER2 was apoptosis accompanied by activation of caspases.


Assuntos
Anticorpos/metabolismo , Apoptose , Ouro/química , Melanoma/patologia , Nanopartículas Metálicas/química , Receptor ErbB-2/metabolismo , Actinas/metabolismo , Fator de Indução de Apoptose/metabolismo , Ciclo Celular , Linhagem Celular Tumoral , Proliferação de Células , Forma Celular , Citocromos c/metabolismo , Proteína-Tirosina Quinases de Adesão Focal/metabolismo , Adesões Focais/metabolismo , Fase G1 , Humanos , Nanopartículas Metálicas/ultraestrutura
19.
Iran J Allergy Asthma Immunol ; 18(2): 182-189, 2019 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-31066254

RESUMO

Systemic sclerosis is a fibrotic autoimmune disease in which aberrant remodeling of the extracellular matrix in organs disturbs their functionalities. The aim of this study was to investigate the expression of gelatinases on systemic sclerosis. Consequently, a mouse model of systemic sclerosis was employed and the gelatinolytic activity of gelatinases was evaluated on the fibrotic tissues of this model. Two groups of ten mice were considered in this work: a group of systemic sclerosis model and control group. For the generation of systemic sclerosis model, mice received bleomycin, while the control group was subjected to phosphate buffered saline (PBS) reception. Mice were tested for fibrosis by using trichrome staining, hydroxyproline measurement and α-SMA detection in tissue sections. Additionally, the gelatinolytic activity of matrix metalloproteinase 2 and matrix metalloproteinase 9 were measured using gelatin zymography in lungs and skin tissue homogenates. The obtained results indicated that subcutaneous injection of bleomycin-induced fibrosis in skin and lung tissues of mice. Pro and active forms of matrix methaloproteinase 9 were increased in fibrotic lung tissues (p<0.05 and p<0.01, respectively), while, the gelatinolytic activity of MMP2 was unaffected in these tissues. Additionally, in skin tissues of bleomycin-treated animals, both pro and active forms of MMP9 and MMP2 were increased (p<0.05). Pro and active forms of gelatinases increase differently in skin and lung tissues of bleomycin-induced scleroderma.


Assuntos
Gelatinases/metabolismo , Escleroderma Sistêmico/metabolismo , Pele/patologia , Actinas/metabolismo , Animais , Bleomicina , Modelos Animais de Doenças , Feminino , Fibrose , Humanos , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Escleroderma Sistêmico/genética , Regulação para Cima
20.
Cell Mol Life Sci ; 76(15): 2933-2956, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31101937

RESUMO

Arginine methylation is a ubiquitous post-translational modification. Three predominant types of arginine-guanidino methylation occur in Eukarya: mono (Rme1/MMA), symmetric (Rme2s/SDMA), and asymmetric (Rme2a/ADMA). Arginine methylation frequently occurs at sites of protein-protein and protein-nucleic acid interactions, providing specificity for binding partners and stabilization of important biological interactions in diverse cellular processes. Each methylarginine isoform-catalyzed by members of the protein arginine methyltransferase family, Type I (PRMT1-4,6,8) and Type II (PRMT5,9)-has unique downstream consequences. Methylarginines are found in ordered domains, domains of low complexity, and in intrinsically disordered regions of proteins-the latter two of which are intimately connected with biological liquid-liquid phase separation. This review highlights discoveries illuminating how arginine methylation affects genome integrity, gene transcription, mRNA splicing and mRNP biology, protein translation and stability, and phase separation. As more proteins and processes are found to be regulated by arginine methylation, its importance for understanding cellular physiology will continue to grow.


Assuntos
Arginina/metabolismo , Proteína-Arginina N-Metiltransferases/metabolismo , Actinas/metabolismo , Cromatina/metabolismo , Reparo do DNA , Histonas/metabolismo , Humanos , Metilação , Processamento de RNA , Ribonucleoproteínas/metabolismo
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