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1.
Mol Biol (Mosk) ; 53(5): 830-837, 2019.
Artigo em Russo | MEDLINE | ID: mdl-31661481

RESUMO

Caspase-8 performs initiatory functions during the induction of apoptosis through the extrinsic pathway. Apoptosis is a type of programmed cell death that plays an important role in regulating embryogenesis and maintaining homeostasis in the tissue of an adult organism, as well as differentiating and removing damaged cells. Dysregulation of the apoptosis mechanisms is associated with the pathogenesis and progression of a number of oncological and neurodegenerative diseases. Caspase-8 (also called СAP4, FLICE, MACH, MCH5) is one of two members of the death effector domain (DED)-containing caspases. Despite the fact that the role of caspase-8 in apoptosis has been well known since the mid 1990s, we are only now beginning to understand the subtle mechanisms of its activation and regulation in response to the activation of death receptors (DRs). In particular, it was demonstrated that the activation of caspase-8 requires the formation of specific oligomeric structures, which are named DED filaments. In this review, the recent data on the mechanisms of activating initiator caspase-8 in DED filaments are considered that allow us to better understand the subtle mechanisms of the initiation of the programmed cell death.


Assuntos
Apoptose , Caspase 8/metabolismo , Precursores Enzimáticos/metabolismo , Citoesqueleto/metabolismo , Ativação Enzimática , Humanos
2.
Adv Exp Med Biol ; 1178: 227-245, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31493230

RESUMO

The cytoskeleton consists of filamentous protein polymers that form organized structures, contributing to a multitude of cell life aspects. It includes three types of polymers: the actin microfilaments, the microtubules and the intermediate filaments. Decades of research have implicated the cytoskeleton in processes that regulate cellular and organismal aging, as well as neurodegeneration associated with injury or neurodegenerative disease, such as Alzheimer's disease, Parkinson's disease, Huntington's disease, Amyotrophic Lateral Sclerosis, or Charcot Marie Tooth disease. Here, we provide a brief overview of cytoskeletal structure and function, and discuss experimental evidence linking cytoskeletal function and dynamics with aging and neurodegeneration.


Assuntos
Envelhecimento , Citoesqueleto , Doenças Neurodegenerativas , Envelhecimento/patologia , Citoesqueleto/química , Citoesqueleto/metabolismo , Humanos , Doenças Neurodegenerativas/patologia
3.
Int J Nanomedicine ; 14: 6197-6215, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31496681

RESUMO

Introduction and objective: Degradation of the extracellular matrix (ECM) changes the physicochemical properties and dysregulates ECM-cell interactions, leading to several pathological conditions, such as invasive cancer. Carbon nanofilm, as a biocompatible and easy to functionalize material, could be used to mimic ECM structures, changing cancer cell behavior to perform like normal cells. Methods: Experiments were performed in vitro with HS-5 cells (as a control) and HepG2 and C3A cancer cells. An aqueous solution of fullerene C60 was used to form a nanofilm. The morphological properties of cells cultivated on C60 nanofilms were evaluated with light, confocal, electron and atomic force microscopy. The cell viability and proliferation were measured by XTT and BrdU assays. Immunoblotting and flow cytometry were used to evaluate the expression level of proliferating cell nuclear antigen and determine the number of cells in the G2/M phase. Results: All cell lines were spread on C60 nanofilms, showing a high affinity to the nanofilm surface. We found that C60 nanofilm mimicked the niche/ECM of cells, was biocompatible and non-toxic, but the mechanical signal from C60 nanofilm created an environment that affected the cell cycle and reduced cell proliferation. Conclusion: The results indicate that C60 nanofilms might be a suitable, substitute component for the niche of cancer cells. The incorporation of fullerene C60 in the ECM/niche may be an alternative treatment for hepatocellular carcinoma.


Assuntos
Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Divisão Celular/efeitos dos fármacos , Fulerenos/farmacologia , Fase G2/efeitos dos fármacos , Neoplasias Hepáticas/patologia , Mecanotransdução Celular , Nanopartículas/química , Comunicação Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Forma Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Citoesqueleto/efeitos dos fármacos , Citoesqueleto/metabolismo , Módulo de Elasticidade , Matriz Extracelular/efeitos dos fármacos , Matriz Extracelular/metabolismo , Fulerenos/química , Humanos , Integrina alfa5beta1/metabolismo , Neoplasias Hepáticas/ultraestrutura , Mecanotransdução Celular/efeitos dos fármacos , Nanopartículas/ultraestrutura , Proteínas de Neoplasias/metabolismo , Transdução de Sinais/efeitos dos fármacos
4.
Gene ; 719: 144074, 2019 Nov 30.
Artigo em Inglês | MEDLINE | ID: mdl-31446094

RESUMO

Kinesin-14 motor es-kifc1 is highly expressed in the male reproductive system of the Chinese mitten crab Eriocheir sinensis (E. sinensis). In addition to acrosomal formation, es-KIFC1 also tightly surrounds the nucleus and its specific mechanism remains unknown. During spermatogenesis, sperm nucleus dents into a cup-shaped structure with several radial arms and completed the nuclear decondensation. In this study, the spatial expression pattern of es-KIFC1 indicates a potential function in nuclear formation with the nuclear localization sequence (NLS) on N-terminal domain which is crucial for the translocation of es-KIFC1 into the nucleus. The Motor domain is associated with microtubule modulation and the Golgi vesicles positioning. Furthermore, the expression level of es-KIFC1 is not only related to the seasonal variation of crustacean development, but also associates with mature sperm storage. The double strand RNA (dsRNA) mediated RNA interference manifests that the cup-shaped sperm nucleus is remarkably malformed and even separates the chromatin throughout the nuclei at the last stage of spermiogenesis. Besides, the sperm nucleus almost disperses its structure and separates the chromatin into several segments throughout the nucleus showing an asymmetrical performance without cytoskeleton. In summary, these results indicate the importance of es-KIFC1 in microtubule positioning and the maintenance of the mature sperm nuclei.


Assuntos
Proteínas de Artrópodes/genética , Braquiúros/fisiologia , Núcleo Celular/metabolismo , Cinesina/genética , Espermatogênese/genética , Animais , Proteínas de Artrópodes/metabolismo , Núcleo Celular/ultraestrutura , Citoesqueleto/metabolismo , Cinesina/metabolismo , Masculino , Microtúbulos/metabolismo , Transporte Proteico , RNA de Cadeia Dupla/genética , Espermatozoides/ultraestrutura
5.
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
6.
Results Probl Cell Differ ; 67: 201-221, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31435796

RESUMO

Organelle positioning as many other morphological parameters in a cell is not random. Centriole positioning as centrosomes or ciliary basal bodies is not an exception to this rule in cell biology. Indeed, centriole positioning is a tightly regulated process that occurs during development, and it is critical for many organs to function properly, not just during development but also in the adulthood. In this book chapter, we overview our knowledge on centriole positioning in different and highly specialized animal cells like photoreceptor or ependymal cells. We will also discuss recent advances in the discovery of molecular pathways involved in this process, mostly related to the cytoskeleton and the cell polarity pathways. And finally, we present quantitative methods that have been used to assess centriole positioning in different cell types although mostly in epithelial cells.


Assuntos
Compartimento Celular , Centríolos/metabolismo , Animais , Polaridade Celular , Citoesqueleto/metabolismo
7.
Int J Nanomedicine ; 14: 3297-3309, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31190794

RESUMO

Background: Cardiovascular disease (CVD) is the leading cause of mortality all over the world. Vascular stents are used to ameliorate vascular stenosis and recover vascular function. The application of nanotubular coatings has been confirmed to promote endothelial cell (EC) proliferation and function. However, the regulatory mechanisms involved in cellular responses to the nanotubular topography have not been defined. In the present study, a microarray analysis was performed to explore the expression patterns of long noncoding RNAs (lncRNAs) in human coronary artery endothelial cells (HCAECs) that were differentially expressed in response to nitinol-based nanotubular coatings. Materials and methods: First, anodization was performed to synthesize nitinol-based nanotubular coatings. Then, HCAECs were cultured on the samples for 24 h to evaluate cell cytoskeleton organization. Next, total RNA was extracted and synthesized into cRNA, which was hybridized onto the microarray. GO analysis and KEGG pathway analysis were performed to investigate the roles of differentially expressed messenger RNAs (mRNAs). Quantitative real-time reverse-transcription polymerase chain reaction (qRT-PCR) was performed to validate the expression of randomly selected lncRNAs. Coexpression networks were created to identify the interactions among lncRNAs and the protein-coding genes involved in nanotubular topography-induced biological and molecular pathways. Independent Student's t-test was applied for comparisons between two groups with statistical significance set at p<0.05. Results: 1085 lncRNAs and 227 mRNAs were significantly differentially expressed in the nitinol-based nanotubular coating group. Bioinformatics analysis revealed that extracellular matrix receptor interactions and cell adhesion molecules play critical roles in the sensing of nitinol-based nanotubular coatings by HCAECs. The TATA-binding protein (TBP) and TBP-associated transfactor 1 (TAF1) are important molecules in EC responses to substrate topography. Conclusion: This study suggests that nanotubular substrate topography regulates ECs by differentially expressed lncRNAs involved extracellular matrix receptor interactions and cell adhesion molecules.


Assuntos
Ligas/farmacologia , Materiais Revestidos Biocompatíveis/farmacologia , Vasos Coronários/citologia , Células Endoteliais/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Nanotubos/química , RNA Longo não Codificante/genética , Proliferação de Células/efeitos dos fármacos , Citoesqueleto/efeitos dos fármacos , Citoesqueleto/metabolismo , Células Endoteliais/efeitos dos fármacos , Perfilação da Expressão Gênica , Ontologia Genética , Redes Reguladoras de Genes/efeitos dos fármacos , Humanos , MicroRNAs/genética , MicroRNAs/metabolismo , Análise em Microsséries , Nanotubos/ultraestrutura , Fenótipo , RNA Longo não Codificante/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Fatores de Transcrição/metabolismo
8.
Emerg Microbes Infect ; 8(1): 934-945, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31237474

RESUMO

Cytoskeletal rearrangement and acute cytotoxicity occur in Vibrio vulnificus-infected host cells. RtxA1 toxin, a multifunctional autoprocessing repeats-in-toxin (MARTX), is essential for the pathogenesis of V. vulnificus and the programmed necrotic cell death. In this study, HeLa cells expressing RtxA1 amino acids 1491-1971 fused to GFP were observed to be rounded. Through yeast two-hybrid screening and subsequent immunoprecipitation validation assays, we confirmed the specific binding of a RtxA11491-1971 fragment with host-cell filamin A, an actin cross-linking scaffold protein. Downregulation of filamin A expression decreased the cytotoxicity of RtxA1 toward host cells. Furthermore, the phosphorylation of JNK and p38 MAPKs was induced by the RtxA1-filamin A interaction during the toxin-mediated cell death. However, the phosphorylation of these MAPKs was not observed during the RtxA1 intoxication of filamin A-deficient M2 cells. In addition, the depletion of pak1, which appeared to be activated by the RtxA1-filamin A interaction, inhibited RtxA1-induced phosphorylation of JNK and p38, and the cells treated with a pak1 inhibitor exhibited decreased RtxA1-mediated cytoskeletal rearrangement and cytotoxicity. Thus, the binding of filamin A by the RtxA11491-1971 domain appears to be a requisite to pak1-mediated MAPK activation, which contributes to the cytoskeletal reorganization and host cell death.


Assuntos
Toxinas Bacterianas/metabolismo , Citoesqueleto/metabolismo , Filaminas/metabolismo , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Vibrioses/metabolismo , Vibrio vulnificus/metabolismo , Quinases Ativadas por p21/metabolismo , Motivos de Aminoácidos , Toxinas Bacterianas/toxicidade , Morte Celular , Citoesqueleto/genética , Filaminas/genética , Células HeLa , Interações Hospedeiro-Patógeno , Humanos , Quinases de Proteína Quinase Ativadas por Mitógeno/genética , Ligação Proteica , Vibrioses/genética , Vibrioses/microbiologia , Vibrioses/fisiopatologia , Vibrio vulnificus/química , Vibrio vulnificus/genética , Quinases Ativadas por p21/genética
9.
Anim Sci J ; 90(7): 801-807, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31134719

RESUMO

Skeletal muscle consists of bundles of myofibers containing millions of myofibrils, each of which is formed of longitudinally aligned sarcomere structures. Sarcomeres are the minimum contractile unit, which mainly consists of four components: Z-bands, thin filaments, thick filaments, and connectin/titin. The size and shape of the sarcomere component is strictly controlled. Surprisingly, skeletal muscle cells not only synthesize a series of myofibrillar proteins but also regulate the assembly of those proteins into the sarcomere structures. However, authentic sarcomere structures cannot be reconstituted by combining purified myofibrillar proteins in vitro, therefore there must be an elaborate mechanism ensuring the correct formation of myofibril structure in skeletal muscle cells. This review discusses the role of myosin, a main component of the thick filament, in thick filament formation and the dynamics of myosin in skeletal muscle cells. Changes in the number of myofibrils in myofibers can cause muscle hypertrophy or atrophy. Therefore, it is important to understand the fundamental mechanisms by which myofibers control myofibril formation at the molecular level to develop approaches that effectively enhance muscle growth in animals.


Assuntos
Citoesqueleto/metabolismo , Músculo Esquelético/citologia , Músculo Esquelético/metabolismo , Miosinas/fisiologia , Animais , Atrofia , Hipertrofia , Músculo Esquelético/crescimento & desenvolvimento , Músculo Esquelético/patologia , Miofibrilas/metabolismo , Miofibrilas/patologia , Miosinas/metabolismo , Sarcômeros/metabolismo
10.
Int J Mol Sci ; 20(9)2019 May 13.
Artigo em Inglês | MEDLINE | ID: mdl-31086053

RESUMO

Calcium-binding proteins (CBPs) can influence and react to Ca2+ transients and modulate the activity of proteins involved in both maintaining homeostatic conditions and protecting cells in harsh environmental conditions. Hibernation is a strategy that evolved in vertebrate and invertebrate species to survive in cold environments; it relies on molecular, cellular, and behavioral adaptations guided by the neuroendocrine system that together ensure unmatched tolerance to hypothermia, hypometabolism, and hypoxia. Therefore, hibernation is a useful model to study molecular neuroprotective adaptations to extreme conditions, and can reveal useful applications to human pathological conditions. In this review, we describe the known changes in Ca2+-signaling and the detection and activity of CBPs in the nervous system of vertebrate and invertebrate models during hibernation, focusing on cytosolic Ca2+ buffers and calmodulin. Then, we discuss these findings in the context of the neuroprotective and neural plasticity mechanisms in the central nervous system: in particular, those associated with cytoskeletal proteins. Finally, we compare the expression of CBPs in the hibernating nervous system with two different conditions of neurodegeneration, i.e., platinum-induced neurotoxicity and Alzheimer's disease, to highlight the similarities and differences and demonstrate the potential of hibernation to shed light into part of the molecular mechanisms behind neurodegenerative diseases.


Assuntos
Proteínas de Ligação ao Cálcio/metabolismo , Sistema Nervoso Central/metabolismo , Sistema Nervoso Central/fisiologia , Hibernação/fisiologia , Neuroproteção/fisiologia , Animais , Citoesqueleto/metabolismo , Humanos
11.
Plant Cell Physiol ; 60(8): 1855-1870, 2019 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-31135031

RESUMO

Formins are evolutionarily conserved eukaryotic proteins engaged in actin nucleation and other aspects of cytoskeletal organization. Angiosperms have two formin clades with multiple paralogs; typical plant Class I formins are integral membrane proteins that can anchor cytoskeletal structures to membranes. For the main Arabidopsis housekeeping Class I formin, FH1 (At3g25500), plasmalemma localization was documented in heterologous expression and overexpression studies. We previously showed that loss of FH1 function increases cotyledon epidermal pavement cell shape complexity via modification of actin and microtubule organization and dynamics. Here, we employ transgenic Arabidopsis expressing green fluorescent protein-tagged FH1 (FH1-GFP) from its native promoter to investigate in vivo behavior of this formin using advanced microscopy techniques. The fusion protein is functional, since its expression complements the fh1 loss-of-function mutant phenotype. Accidental overexpression of FH1-GFP results in a decrease in trichome branch number, while fh1 mutation has the opposite effect, indicating a general role of this formin in controlling cell shape complexity. Consistent with previous reports, FH1-GFP associates with membranes. However, the protein exhibits surprising actin- and secretory pathway-dependent dynamic localization and relocates between cellular endomembranes and the plasmalemma during cell division and differentiation in root tissues, with transient tonoplast localization at the transition/elongation zones border. FH1-GFP also accumulates in actin-rich regions of cortical cytoplasm and associates with plasmodesmata in both the cotyledon epidermis and root tissues. Together with previous reports from metazoan systems, this suggests that formins might have a shared (ancestral or convergent) role at cell-cell junctions.


Assuntos
Arabidopsis/metabolismo , Raízes de Plantas/metabolismo , Plasmodesmos/fisiologia , Arabidopsis/citologia , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Citoesqueleto/genética , Citoesqueleto/metabolismo , Raízes de Plantas/citologia
12.
Cell Mol Life Sci ; 76(18): 3571-3581, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31143959

RESUMO

Apoptosis plays a crucial role in clearing old or critically compromised cells, and actively maintains epithelial homeostasis and epithelial morphogenesis during embryo development. But how is the apoptotic signaling pathway able to orchestrate such complex and dynamic multi-cellular morphological events at the tissue scale? In this review we collected the most updated knowledge regarding how apoptosis controls different cytoskeletal components. We describe how apoptosis can control epithelial homeostasis though epithelial extrusion, a highly orchestrated process based on high- order actomyosin structures and on the coordination between the apoptotic and the neighboring cells. Finally, we describe how the synergy among forces generated by multiple apoptotic cells can shape epithelia in embryo development.


Assuntos
Apoptose , Células Epiteliais/metabolismo , Transdução de Sinais/fisiologia , Animais , Citoesqueleto/metabolismo , Desenvolvimento Embrionário , Células Epiteliais/citologia , Homeostase , Miotonina Proteína Quinase/metabolismo , Quinases Associadas a rho/metabolismo
13.
Ecotoxicol Environ Saf ; 180: 269-279, 2019 Sep 30.
Artigo em Inglês | MEDLINE | ID: mdl-31100591

RESUMO

With the broad application of nanoparticles, nanotoxicology has attracted substantial attention in environmental science. However, the methods for detecting few and targeted genes or proteins, even single omics approaches, may miss other responses, including the major responses induced by nanoparticles. To determine the actual toxicological mechanisms of zebrafish brains induced by graphene oxide (GO, a popular carbon-based nanomaterial applied in various fields) at nonlethal concentrations, multi-omics and regular analyses were combined. The biomolecule responses were remarkable, although GO was not obviously observed in brain tissues. The trends for gene and protein changes were the same and accounted for 3.53% and 5.36% of all changes in the genome and proteome, respectively, suggesting a limitation of single omics analysis. Transcriptomics and proteomics analyses indicated that GO affected the functions or pathways of the troponin complex, actin cytoskeleton, monosaccharide transmembrane transporter activity, oxidoreductase activity and focal adhesion. Both metabolomics and proteomics revealed mitochondrial dysfunction and disruption of the citric acid cycle. The integrated analysis of omics, transmission electron microscopy and immunostaining confirmed that GO induced energy disruptions and mitochondrial damage by downregulating tubulin. The combination of multi-omics and regular analyses provides insights into the actual and highly influential mechanisms underlying nanotoxicity.


Assuntos
Encéfalo/efeitos dos fármacos , Grafite/toxicidade , Nanopartículas/toxicidade , Poluentes Químicos da Água/toxicidade , Peixe-Zebra/metabolismo , Animais , Encéfalo/metabolismo , Citoesqueleto/metabolismo , Relação Dose-Resposta a Droga , Metabolismo Energético/efeitos dos fármacos , Perfilação da Expressão Gênica/métodos , Metabolômica/métodos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Proteômica/métodos , Peixe-Zebra/genética
14.
Int J Mol Sci ; 20(9)2019 Apr 30.
Artigo em Inglês | MEDLINE | ID: mdl-31052288

RESUMO

A critical aim in neuroscience is to obtain a comprehensive view of how regulated neurotransmission is achieved. Our current understanding of synapses relies mainly on data from electrophysiological recordings, imaging, and molecular biology. Based on these methodologies, proteins involved in a synaptic vesicle (SV) formation, mobility, and fusion at the active zone (AZ) membrane have been identified. In the last decade, electron tomography (ET) combined with a rapid freezing immobilization of neuronal samples opened a window for understanding the structural machinery with the highest spatial resolution in situ. ET provides significant insights into the molecular architecture of the AZ and the organelles within the presynaptic nerve terminal. The specialized sensory ribbon synapses exhibit a distinct architecture from neuronal synapses due to the presence of the electron-dense synaptic ribbon. However, both synapse types share the filamentous structures, also commonly termed as tethers that are proposed to contribute to different steps of SV recruitment and exocytosis. In this review, we discuss the emerging views on the role of filamentous structures in SV exocytosis gained from ultrastructural studies of excitatory, mainly central neuronal compared to ribbon-type synapses with a focus on inner hair cell (IHC) ribbon synapses. Moreover, we will speculate on the molecular entities that may be involved in filament formation and hence play a crucial role in the SV cycle.


Assuntos
Citoesqueleto/metabolismo , Exocitose , Células Receptoras Sensoriais/metabolismo , Membranas Sinápticas/metabolismo , Vesículas Sinápticas/metabolismo , Animais , Citoesqueleto/ultraestrutura , Humanos , Células Receptoras Sensoriais/ultraestrutura , Membranas Sinápticas/ultraestrutura , Transmissão Sináptica , Vesículas Sinápticas/ultraestrutura
15.
Int J Mol Sci ; 20(10)2019 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-31096581

RESUMO

The FLUMIAS (Fluorescence-Microscopic Analyses System for Life-Cell-Imaging in Space) confocal laser spinning disk fluorescence microscope represents a new imaging capability for live cell imaging experiments on suborbital ballistic rocket missions. During the second pioneer mission of this microscope system on the TEXUS-54 suborbital rocket flight, we developed and performed a live imaging experiment with primary human macrophages. We simultaneously imaged four different cellular structures (nucleus, cytoplasm, lysosomes, actin cytoskeleton) by using four different live cell dyes (Nuclear Violet, Calcein, LysoBrite, SiR-actin) and laser wavelengths (405, 488, 561, and 642 nm), and investigated the cellular morphology in microgravity (10-4 to 10-5 g) over a period of about six minutes compared to 1 g controls. For live imaging of the cytoskeleton during spaceflight, we combined confocal laser microscopy with the SiR-actin probe, a fluorogenic silicon-rhodamine (SiR) conjugated jasplakinolide probe that binds to F-actin and displays minimal toxicity. We determined changes in 3D cell volume and surface, nuclear volume and in the actin cytoskeleton, which responded rapidly to the microgravity environment with a significant reduction of SiR-actin fluorescence after 4-19 s microgravity, and adapted subsequently until 126-151 s microgravity. We conclude that microgravity induces geometric cellular changes and rapid response and adaptation of the potential gravity-transducing cytoskeleton in primary human macrophages.


Assuntos
Citoesqueleto/metabolismo , Macrófagos/citologia , Macrófagos/metabolismo , Ausência de Peso , Citoesqueleto de Actina , Actinas/metabolismo , Linhagem Celular , Núcleo Celular , Citoplasma , Humanos , Lisossomos , Microscopia Confocal/instrumentação , Microscopia Confocal/métodos , Microscopia de Fluorescência/instrumentação , Microscopia de Fluorescência/métodos , Monócitos/citologia , Voo Espacial
16.
Parasitol Res ; 118(6): 1899-1918, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30949853

RESUMO

After host cell invasion, Toxoplasma secretes a variety of dense granule proteins (GRA proteins) from its secretory dense granules, which are involved in the biogenesis of the parasitophorous vacuole (PV). TgGRA8I is predicted to contain proline-rich domains, which are structural features of some cytoskeleton-related proteins. In agreement with this observation, previous proteomic analyses revealed the presence of TgGRA8I in the Toxoplasma sub-pellicular cytoskeleton. In the present study, we show (1) by docking analyses that TgGRA8I may interact with both Toxoplasma ß-tubulin and actin; (2) by immunoelectron microscopy, proteomic, biochemical, and cellular approaches that TgGRA8I associates with sub-pellicular microtubules and actin at the parasite sub-pellicular cytoskeleton; (3) that type I parasites (RH strain) lacking the GRA8 gene (RHΔku80Δgra8) exhibit loss of conoid extrusion, diminished cell infection, and egress capabilities, and that these motility impairments were likely due to important alterations in their sub-pellicular cytoskeleton, in particular their sub-pellicular microtubules and meshwork. Parasites lacking the GRA4 gene (RHΔku80Δgra4) did not show modifications in the organization of the sub-pellicular cytoskeleton. Collectively, these results demonstrated that TgGRA8I is a dense granule protein that, besides its role in the formation of the PV, contributes to the organization of the parasite sub-pellicular cytoskeleton and motility. This is the first proline-rich protein described in the Toxoplasma cytoskeleton, which is a key organelle for both the parasite motility and the invasion process. Knowledge about the function of cytoskeleton components in Toxoplasma is fundamental to understand the motility process and the host cell invasion mechanism. Refining this knowledge should lead to the design of novel pharmacological strategies for the treatment against toxoplasmosis.


Assuntos
Actinas/metabolismo , Antígenos de Protozoários/metabolismo , Movimento Celular/genética , Citoesqueleto/metabolismo , Proteínas de Protozoários/metabolismo , Toxoplasma/metabolismo , Toxoplasma/patogenicidade , Tubulina (Proteína)/metabolismo , Animais , Antígenos de Protozoários/genética , Transporte Biológico , Microscopia Imunoeletrônica , Microtúbulos/metabolismo , Simulação de Acoplamento Molecular , Proteômica , Proteínas de Protozoários/genética , Vesículas Secretórias/metabolismo , Toxoplasma/genética , Toxoplasmose/parasitologia , Toxoplasmose/patologia , Vacúolos/parasitologia
17.
Biomed Res ; 40(2): 79-85, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30982803

RESUMO

The vitreous of perinatal mice temporarily develops a unique vascular system, called the vasa hyaloidea propria (VHP). Observations showed the vessels possessed an extracellular matrix including the basement membrane in their entire length. Immunostaining of whole mount preparations of VHP with integrin ß1 antibody displayed a bush-like network consisting of long and straight fibers which were associated with the VHP but extended apart from the blood vessels. Electron microscopically, each fiber was composed of a bundle of thin filaments different from collagen fibrils. Macrophages associated with the VHP appeared to be arrested by the integrin bushes. The integrin bushes fragmented and disappeared by postnatal day 10, just before the regression of the VHP. Macrophages were involved in the digestion and clearance of integrin bushes. The vitreous integrin bushes appear to provide a scaffold for architectural maintenance of the hyaloid vessels and macrophages.


Assuntos
Membrana Basal/ultraestrutura , Vasos Sanguíneos/ultraestrutura , Citoesqueleto/ultraestrutura , Matriz Extracelular/ultraestrutura , Integrina beta1/ultraestrutura , Corpo Vítreo/ultraestrutura , Animais , Animais Recém-Nascidos , Membrana Basal/metabolismo , Vasos Sanguíneos/anatomia & histologia , Citoesqueleto/metabolismo , Embrião de Mamíferos , Matriz Extracelular/metabolismo , Feminino , Expressão Gênica , Imuno-Histoquímica , Integrina beta1/genética , Integrina beta1/metabolismo , Macrófagos/metabolismo , Macrófagos/ultraestrutura , Camundongos , Microscopia Eletrônica , Neovascularização Fisiológica , Gravidez , Corpo Vítreo/anatomia & histologia , Corpo Vítreo/irrigação sanguínea
18.
Nat Cell Biol ; 21(5): 603-613, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30988424

RESUMO

Mitochondrial fission involves the preconstriction of an organelle followed by scission by dynamin-related protein Drp1. Preconstriction is facilitated by actin and non-muscle myosin II through a mechanism that remains unclear, largely due to the unknown cytoskeletal ultrastructure at mitochondrial constrictions. Here, using platinum replica electron microscopy, we show that mitochondria in cells are embedded in an interstitial cytoskeletal network that contains abundant unbranched actin filaments. Both spontaneous and induced mitochondrial constrictions typically associate with a criss-cross array of long actin filaments that comprise part of this interstitial network. Non-muscle myosin II is found adjacent to mitochondria but is not specifically enriched at the constriction sites. During ionomycin-induced mitochondrial fission, F-actin clouds colocalize with mitochondrial constriction sites, whereas dynamic myosin II clouds are present in the vicinity of constrictions. We propose that myosin II promotes mitochondrial constriction by inducing stochastic deformations of the interstitial actin network, which applies pressure on the mitochondrial surface and thus initiates curvature-sensing mechanisms that complete mitochondrial constriction.


Assuntos
Actinas/genética , Citoesqueleto/ultraestrutura , Mitocôndrias/ultraestrutura , Dinâmica Mitocondrial/genética , Miosina Tipo II/genética , Citoesqueleto de Actina/química , Citoesqueleto de Actina/ultraestrutura , Actinas/metabolismo , Animais , Células COS , Cercopithecus aethiops , Constrição , Citoesqueleto/metabolismo , Ionomicina/farmacologia , Mitocôndrias/genética , Dinâmica Mitocondrial/efeitos dos fármacos , Proteínas Mitocondriais/química , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , Miosina Tipo II/química , Miosina Tipo II/metabolismo
19.
Int J Mol Sci ; 20(7)2019 Mar 29.
Artigo em Inglês | MEDLINE | ID: mdl-30934860

RESUMO

The essential role of Hippo signaling pathway in cancer development has been elucidated by recent studies. In the gastrointestinal tissues, deregulation of the Hippo pathway is one of the most important driving events for tumorigenesis. It is widely known that Yes-associated protein 1 (YAP1) and WW domain that contain transcription regulator 1 (TAZ), two transcriptional co-activators with a PDZ-binding motif, function as critical effectors negatively regulated by the Hippo pathway. Previous studies indicate the involvement of YAP1/TAZ in mechanotransduction by crosstalking with the extracellular matrix (ECM) and the F-actin cytoskeleton associated signaling network. In gastric cancer (GC), YAP1/TAZ functions as an oncogene and transcriptionally promotes tumor formation by cooperating with TEAD transcription factors. Apart from the classic role of Hippo-YAP1 cascade, in this review, we summarize the current investigations to highlight the prominent role of YAP1/TAZ as a mechanical sensor and responder under mechanical stress and address its potential prognostic and therapeutic value in GC.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Carcinogênese/metabolismo , Carcinogênese/patologia , Citoesqueleto/metabolismo , Mecanotransdução Celular , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patologia , Animais , Humanos , Modelos Biológicos
20.
Nat Commun ; 10(1): 1766, 2019 04 16.
Artigo em Inglês | MEDLINE | ID: mdl-30992433

RESUMO

The single nucleotide polymorphism (SNP) rs744373 in the bridging integrator-1 gene (BIN1) is a risk factor for Alzheimer's disease (AD). In the brain, BIN1 is involved in endocytosis and sustaining cytoskeleton integrity. Post-mortem and in vitro studies suggest that BIN1-associated AD risk is mediated by increased tau pathology but whether rs744373 is associated with increased tau pathology in vivo is unknown. Here we find in 89 older individuals without dementia, that BIN1 rs744373 risk-allele carriers show higher AV1451 tau-PET across brain regions corresponding to Braak stages II-VI. In contrast, the BIN1 rs744373 SNP was not associated with AV45 amyloid-PET uptake. Furthermore, the rs744373 risk-allele was associated with worse memory performance, mediated by increased global tau levels. Together, our findings suggest that the BIN1 rs744373 SNP is associated with increased tau but not beta-amyloid pathology, suggesting that alterations in BIN1 may contribute to memory deficits via increased tau pathology.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Doença de Alzheimer/genética , Memória , Proteínas Nucleares/genética , Polimorfismo de Nucleotídeo Único , Proteínas Supressoras de Tumor/genética , Proteínas tau/metabolismo , Idoso , Idoso de 80 Anos ou mais , Alelos , Doença de Alzheimer/metabolismo , Encéfalo/diagnóstico por imagem , Cognição , Citoesqueleto/metabolismo , Endocitose , Feminino , Variação Genética , Humanos , Imagem por Ressonância Magnética , Masculino , Tomografia por Emissão de Pósitrons , Fatores de Risco
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