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1.
Nat Commun ; 10(1): 4770, 2019 10 18.
Artigo em Inglês | MEDLINE | ID: mdl-31628317

RESUMO

Animals perform or terminate particular behaviors by integrating external cues and internal states through neural circuits. Identifying neural substrates and their molecular modulators promoting or inhibiting animal behaviors are key steps to understand how neural circuits control behaviors. Here, we identify the Cholecystokinin-like peptide Drosulfakinin (DSK) that functions at single-neuron resolution to suppress male sexual behavior in Drosophila. We found that Dsk neurons physiologically interact with male-specific P1 neurons, part of a command center for male sexual behaviors, and function oppositely to regulate multiple arousal-related behaviors including sex, sleep and spontaneous walking. We further found that the DSK-2 peptide functions through its receptor CCKLR-17D3 to suppress sexual behaviors in flies. Such a neuropeptide circuit largely overlaps with the fruitless-expressing neural circuit that governs most aspects of male sexual behaviors. Thus DSK/CCKLR signaling in the sex circuitry functions antagonistically with P1 neurons to balance arousal levels and modulate sexual behaviors.


Assuntos
Proteínas de Drosophila/metabolismo , Drosophila melanogaster/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Neurônios/metabolismo , Neuropeptídeos/metabolismo , Oligopeptídeos/metabolismo , Fatores de Transcrição/metabolismo , Animais , Animais Geneticamente Modificados , Nível de Alerta/fisiologia , Comunicação Celular/genética , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Drosophila melanogaster/fisiologia , Feminino , Locomoção/fisiologia , Masculino , Proteínas do Tecido Nervoso/genética , Neurônios/citologia , Neuropeptídeos/genética , Oligopeptídeos/genética , Comportamento Sexual Animal/fisiologia , Transdução de Sinais/genética , Sono/fisiologia , Fatores de Transcrição/genética
2.
Hypertension ; 74(5): 1152-1159, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31564164

RESUMO

Microarray comparison of the transcriptomes of human adrenal zona glomerulosa (ZG) and zona fasciculata found several ZG-specific genes that negatively regulate aldosterone secretion. The third and most significantly upregulated ZG-gene (19.9-fold compared with zona fasciculata, P=6.58×10-24) was ANO4, a putative Ca2+-activated chloride channel. We have investigated the role of ANO4 in human adrenal, and whether it functions like the prototype anoctamin, ANO1. We evaluated ANO4 mRNA and protein expression in human adrenal by qPCR and immunohistochemistry, compared the effects of ANO4 and ANO1 overexpression on baseline and stimulated aldosterone secretion and cell proliferation in H295R cells, and analyzed ANO4 activity as a Ca2+-activated chloride channel in comparison with other anoctamins by a fluorescence-based functional assay. The expression of ANO4 in ZG was confirmed by qPCR as 23.21-fold upregulated compared with zona fasciculata (n=18; P=4.93×10-7). Immunohistochemistry found cytoplasmic, ZG-selective expression of ANO4 (anoctamin 4) protein. ANO4 overexpression in H295R cells attenuated calcium-mediated aldosterone secretion and cell proliferation in comparison to controls. The latter effects were in a different direction to those of ANO1. The functional assay showed that, in contrast to ANO1, ANO4 expression results in low levels of calcium-dependent anion transport. In conclusion, ANO4 is one of the most highly expressed genes in ZG. It attenuates stimulated aldosterone secretion and cell proliferation. Although belonging to a family of Ca2+-activated chloride channels, it does not generate significant plasma membrane chloride channel activity.


Assuntos
Aldosterona/biossíntese , Anoctaminas/genética , Regulação da Expressão Gênica , Hiperaldosteronismo/genética , Hipertensão/fisiopatologia , Transdução de Sinais/genética , Zona Glomerulosa/metabolismo , Córtex Suprarrenal/citologia , Córtex Suprarrenal/metabolismo , Córtex Suprarrenal/patologia , Análise de Variância , Comunicação Celular/genética , Proliferação de Células , Células Cultivadas , Imunofluorescência , Humanos , Hiperaldosteronismo/patologia , Hipertensão/etiologia , Reação em Cadeia da Polimerase em Tempo Real/métodos , Análise Serial de Tecidos , Técnicas de Cultura de Tecidos , Transcriptoma/genética , Regulação para Cima , Zona Fasciculada/metabolismo , Zona Fasciculada/patologia , Zona Glomerulosa/patologia
3.
Nat Neurosci ; 22(10): 1696-1708, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31551601

RESUMO

The mammalian brain is complex, with multiple cell types performing a variety of diverse functions, but exactly how each cell type is affected in aging remains largely unknown. Here we performed a single-cell transcriptomic analysis of young and old mouse brains. We provide comprehensive datasets of aging-related genes, pathways and ligand-receptor interactions in nearly all brain cell types. Our analysis identified gene signatures that vary in a coordinated manner across cell types and gene sets that are regulated in a cell-type specific manner, even at times in opposite directions. These data reveal that aging, rather than inducing a universal program, drives a distinct transcriptional course in each cell population, and they highlight key molecular processes, including ribosome biogenesis, underlying brain aging. Overall, these large-scale datasets (accessible online at https://portals.broadinstitute.org/single_cell/study/aging-mouse-brain ) provide a resource for the neuroscience community that will facilitate additional discoveries directed towards understanding and modifying the aging process.


Assuntos
Envelhecimento/genética , Encéfalo/crescimento & desenvolvimento , Neurônios/fisiologia , Análise de Célula Única , Transcriptoma/genética , Animais , Encéfalo/citologia , Comunicação Celular/genética , Linhagem da Célula/genética , Perfilação da Expressão Gênica , Regulação da Expressão Gênica/genética , Sequenciamento de Nucleotídeos em Larga Escala , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Ribossomos/genética
4.
Mol Cell ; 75(3): 644-660.e5, 2019 08 08.
Artigo em Inglês | MEDLINE | ID: mdl-31398325

RESUMO

Cell-cell communication via ligand-receptor signaling is a fundamental feature of complex organs. Despite this, the global landscape of intercellular signaling in mammalian liver has not been elucidated. Here we perform single-cell RNA sequencing on non-parenchymal cells isolated from healthy and NASH mouse livers. Secretome gene analysis revealed a highly connected network of intrahepatic signaling and disruption of vascular signaling in NASH. We uncovered the emergence of NASH-associated macrophages (NAMs), which are marked by high expression of triggering receptors expressed on myeloid cells 2 (Trem2), as a feature of mouse and human NASH that is linked to disease severity and highly responsive to pharmacological and dietary interventions. Finally, hepatic stellate cells (HSCs) serve as a hub of intrahepatic signaling via HSC-derived stellakines and their responsiveness to vasoactive hormones. These results provide unprecedented insights into the landscape of intercellular crosstalk and reprogramming of liver cells in health and disease.


Assuntos
Comunicação Celular/genética , Fígado/metabolismo , Hepatopatia Gordurosa não Alcoólica/genética , Análise de Sequência de RNA , Animais , Reprogramação Celular/genética , Modelos Animais de Doenças , Células Estreladas do Fígado/metabolismo , Células Estreladas do Fígado/patologia , Humanos , Ligantes , Fígado/patologia , Macrófagos/metabolismo , Macrófagos/patologia , Camundongos , Hepatopatia Gordurosa não Alcoólica/metabolismo , Hepatopatia Gordurosa não Alcoólica/patologia , Transdução de Sinais/genética , Análise de Célula Única
5.
PLoS Genet ; 15(8): e1008377, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31465456

RESUMO

Intercellular communication in adjacent cell layers determines cell fate and polarity, thus orchestrating tissue specification and differentiation. Here we use the maize stomatal apparatus as a model to investigate cell fate determination. Mutations in ZmBZU2 (bizui2, bzu2) confer a complete absence of subsidiary cells (SCs) and normal guard cells (GCs), leading to failure of formation of mature stomatal complexes. Nuclear polarization and actin accumulation at the interface between subsidiary mother cells (SMCs) and guard mother cells (GMCs), an essential pre-requisite for asymmetric cell division, did not occur in Zmbzu2 mutants. ZmBZU2 encodes a basic helix-loop-helix (bHLH) transcription factor, which is an ortholog of AtMUTE in Arabidopsis (BZU2/ZmMUTE). We found that a number of genes implicated in stomatal development are transcriptionally regulated by BZU2/ZmMUTE. In particular, BZU2/ZmMUTE directly binds to the promoters of PAN1 and PAN2, two early regulators of protodermal cell fate and SMC polarization, consistent with the low levels of transcription of these genes observed in bzu2-1 mutants. BZU2/ZmMUTE has the cell-to-cell mobility characteristic similar to that of BdMUTE in Brachypodium distachyon. Unexpectedly, BZU2/ZmMUTE is expressed in GMC from the asymmetric division stage to the GMC division stage, and especially in the SMC establishment stage. Taken together, these data imply that BZU2/ZmMUTE is required for early events in SMC polarization and differentiation as well as for the last symmetrical division of GMCs to produce the two GCs, and is a master determinant of the cell fate of its neighbors through cell-to-cell communication.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Regulação da Expressão Gênica de Plantas , Proteínas de Plantas/metabolismo , Células-Tronco/fisiologia , Zea mays/fisiologia , Fatores de Transcrição Hélice-Alça-Hélice Básicos/genética , Comunicação Celular/genética , Diferenciação Celular/genética , Divisão Celular/genética , Polaridade Celular/genética , Mutação , Proteínas de Plantas/genética , Estômatos de Plantas/citologia , Estômatos de Plantas/fisiologia , Plantas Geneticamente Modificadas , Regiões Promotoras Genéticas/genética
6.
Biomed Res Int ; 2019: 5013592, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31380426

RESUMO

Cell fusion is a highly regulated biological process that occurs under both physiological and pathological conditions. The cellular and extracellular environment is critical for the induction of the cell-cell fusion. Aberrant cell fusion is initiated during tumor progression. Tumor microenvironment is a complex dynamic system formed by the interaction between tumor cells and their surrounding cells. Cell-cell fusion mediates direct interaction between tumor cells and their surrounding cells and is associated with tumor initiation and progression. Various microenvironmental factors affect cell fusion in tumor microenvironment and generate hybrids that acquire genomes of both parental cells and exhibit novel characteristics, such as tumor stem cell-like properties, radioresistance, drug resistance, immune evasion, and enhanced migration and invasion abilities, which are closely related to the initiation, invasion, and metastasis of tumor. The phenotypic characteristics of hybrids are based on the phenotypes of parental cells, and the fusion of tumor cells with diverse types of microenvironmental fusogenic cells is concomitant with phenotypic heterogeneity. This review highlights the types of fusogenic cells in tumor microenvironment that can fuse with tumor cells and their specific significance and summarizes the various microenvironmental factors affecting tumor cell fusion. This review may be used as a reference to develop strategies for future research on tumor cell fusion and the exploration of cell fusion-based antitumor therapies.


Assuntos
Comunicação Celular/genética , Transformação Celular Neoplásica/genética , Neoplasias/genética , Microambiente Tumoral/genética , Resistencia a Medicamentos Antineoplásicos/genética , Humanos , Metástase Neoplásica , Neoplasias/patologia , Células-Tronco Neoplásicas/metabolismo , Células-Tronco Neoplásicas/patologia , Fenótipo
7.
Int J Mol Sci ; 20(15)2019 Jul 24.
Artigo em Inglês | MEDLINE | ID: mdl-31344783

RESUMO

Investigations of information dynamics in eukaryotic cells focus almost exclusively on heritable information in the genome. Gene networks are modeled as "central processors" that receive, analyze, and respond to intracellular and extracellular signals with the nucleus described as a cell's control center. Here, we present a model in which cellular information is a distributed system that includes non-genomic information processing in the cell membrane that may quantitatively exceed that of the genome. Within this model, the nucleus largely acts a source of macromolecules and processes information needed to synchronize their production with temporal variations in demand. However, the nucleus cannot produce microsecond responses to acute, life-threatening perturbations and cannot spatially resolve incoming signals or direct macromolecules to the cellular regions where they are needed. In contrast, the cell membrane, as the interface with its environment, can rapidly detect, process, and respond to external threats and opportunities through the large amounts of potential information encoded within the transmembrane ion gradient. Our model proposes environmental information is detected by specialized protein gates within ion-specific transmembrane channels. When the gate receives a specific environmental signal, the ion channel opens and the received information is communicated into the cell via flow of a specific ion species (i.e., K+, Na+, Cl-, Ca2+, Mg2+) along electrochemical gradients. The fluctuation of an ion concentration within the cytoplasm adjacent to the membrane channel can elicit an immediate, local response by altering the location and function of peripheral membrane proteins. Signals that affect a larger surface area of the cell membrane and/or persist over a prolonged time period will produce similarly cytoplasmic changes on larger spatial and time scales. We propose that as the amplitude, spatial extent, and duration of changes in cytoplasmic ion concentrations increase, the information can be communicated to the nucleus and other intracellular structure through ion flows along elements of the cytoskeleton to the centrosome (via microtubules) or proteins in the nuclear membrane (via microfilaments). These dynamics add spatial and temporal context to the more well-recognized information communication from the cell membrane to the nucleus following ligand binding to membrane receptors. Here, the signal is transmitted and amplified through transduction by the canonical molecular (e.g., Mitogen Activated Protein Kinases (MAPK) pathways. Cytoplasmic diffusion allows this information to be broadly distributed to intracellular organelles but at the cost of loss of spatial and temporal information also contained in ligand binding.


Assuntos
Comunicação Celular/genética , Membrana Celular/genética , Núcleo Celular/genética , Células Eucarióticas , Cálcio/metabolismo , Citoplasma/genética , Citoesqueleto/genética , Genoma/genética , Canais Iônicos/genética , Canais Iônicos/metabolismo , Íons/metabolismo , Transdução de Sinais/genética
8.
Sensors (Basel) ; 19(15)2019 Jul 24.
Artigo em Inglês | MEDLINE | ID: mdl-31344821

RESUMO

Purinergic signals, such as extracellular adenosine triphosphate (ATP) and adenosine diphosphate (ADP), mediate intercellular communication and stress responses throughout mammalian tissues, but the dynamics of their release and clearance are still not well understood. Although physiochemical methods provide important insight into physiology, genetically encoded optical sensors have proven particularly powerful in the quantification of signaling in live specimens. Indeed, genetically encoded luminescent and fluorescent sensors provide new insights into ATP-mediated purinergic signaling. However, new tools to detect extracellular ADP are still required. To this end, in this study, we use protein engineering to generate a new genetically encoded sensor that employs a high-affinity bacterial ADP-binding protein and reports a change in occupancy with a change in the Förster-type resonance energy transfer (FRET) between cyan and yellow fluorescent proteins. We characterize the sensor in both protein solution studies, as well as live-cell microscopy. This new sensor responds to nanomolar and micromolar concentrations of ADP and ATP in solution, respectively, and in principle it is the first fully-genetically encoded sensor with sufficiently high affinity for ADP to detect low levels of extracellular ADP. Furthermore, we demonstrate that tethering the sensor to the cell surface enables the detection of physiologically relevant nucleotide release induced by hypoosmotic shock as a model of tissue edema. Thus, we provide a new tool to study purinergic signaling that can be used across genetically tractable model systems.


Assuntos
Difosfato de Adenosina/isolamento & purificação , Trifosfato de Adenosina/isolamento & purificação , Técnicas Biossensoriais , Edema/diagnóstico , Difosfato de Adenosina/química , Trifosfato de Adenosina/química , Comunicação Celular/genética , Edema/genética , Transferência Ressonante de Energia de Fluorescência , Corantes Fluorescentes/química , Humanos , Proteínas Luminescentes/química , Pressão Osmótica , Ligação Proteica/genética
9.
Nat Commun ; 10(1): 2481, 2019 06 06.
Artigo em Inglês | MEDLINE | ID: mdl-31171792

RESUMO

Mammary stroma is essential for epithelial morphogenesis and development. Indeed, postnatal mammary gland (MG) development is controlled locally by the repetitive and bi-directional cross-talk between the epithelial and the stromal compartment. However, the signalling pathways involved in stromal-epithelial communication are not entirely understood. Here, we identify Sfrp3 as a mediator of the stromal-epithelial communication that is required for normal mouse MG development. Using Drosophila wing imaginal disc, we demonstrate that Sfrp3 functions as an extracellular transporter of Wnts that facilitates their diffusion, and thus, their levels in the boundaries of different compartments. Indeed, loss of Sfrp3 in mice leads to an increase of ductal invasion and branching mirroring an early pregnancy state. Finally, we observe that loss of Sfrp3 predisposes for invasive breast cancer. Altogether, our study shows that Sfrp3 controls MG morphogenesis by modulating the stromal-epithelial cross-talk during pubertal development.


Assuntos
Comunicação Celular/genética , Células Epiteliais/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/genética , Glândulas Mamárias Animais/crescimento & desenvolvimento , Neoplasias Mamárias Animais/genética , Células Estromais/metabolismo , Proteínas Wnt/metabolismo , Animais , Drosophila , Proteínas de Drosophila , Feminino , Discos Imaginais , Glândulas Mamárias Animais/metabolismo , Camundongos , Camundongos Knockout , Morfogênese , Gravidez , Maturidade Sexual , Fatores de Transcrição , Via de Sinalização Wnt
10.
PLoS Comput Biol ; 15(6): e1007030, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31194728

RESUMO

Prolactin is a major hormone product of the pituitary gland, the central endocrine regulator. Despite its physiological importance, the cell-level mechanisms of prolactin production are not well understood. Having significantly improved the resolution of real-time-single-cell-GFP-imaging, the authors recently revealed that prolactin gene transcription is highly dynamic and stochastic yet shows space-time coordination in an intact tissue slice. However, it still remains an open question as to what kind of cellular communication mediates the observed space-time organization. To determine the type of interaction between cells we developed a statistical model. The degree of similarity between two expression time series was studied in terms of two distance measures, Euclidean and geodesic, the latter being a network-theoretic distance defined to be the minimal number of edges between nodes, and this was used to discriminate between juxtacrine from paracrine signalling. The analysis presented here suggests that juxtacrine signalling dominates. To further determine whether the coupling is coordinating transcription or post-transcriptional activities we used stochastic switch modelling to infer the transcriptional profiles of cells and estimated their similarity measures to deduce that their spatial cellular coordination involves coupling of transcription via juxtacrine signalling. We developed a computational model that involves an inter-cell juxtacrine coupling, yielding simulation results that show space-time coordination in the transcription level that is in agreement with the above analysis. The developed model is expected to serve as the prototype for the further study of tissue-level organised gene expression for epigenetically regulated genes, such as prolactin.


Assuntos
Comunicação Celular/genética , Modelos Biológicos , Comunicação Parácrina/genética , Animais , Comunicação Celular/fisiologia , Biologia Computacional , Regulação da Expressão Gênica/genética , Humanos , Masculino , Comunicação Parácrina/fisiologia , Hipófise/metabolismo , Prolactina/genética , Prolactina/metabolismo , Ratos , Ratos Transgênicos , Processos Estocásticos
11.
Biochim Biophys Acta Rev Cancer ; 1872(1): 89-102, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31202687

RESUMO

Metastasis is a complex systemic disease that develops as a result of interactions between tumor cells and their local and distant microenvironments. Local and systemic immune-related changes play especially critical roles in limiting or enabling the development of metastatic disease. Although anti-tumor immune responses likely eliminate most early primary and metastatic lesions, factors secreted by cancer or stromal cells in the primary tumor can mobilize and activate cells in distant organs in a way that promotes the outgrowth of disseminated cancer cells into macrometastatic lesions. Therefore, the prevention, detection, and effective treatment of metastatic disease require a deeper understanding of the systemic effects of primary tumors as well as predisposing hereditary and acquired host factors including chronic inflammatory conditions. The success of immunotherapy in a subset of cancer patients is an example of how modulating the microenvironment and tumor-immune cell interactions can be exploited for the effective eradiation of even advanced-stage tumors. Here, we highlight emerging insights and clinical implications of cancer as a systemic disease.


Assuntos
Imunoterapia , Inflamação/genética , Metástase Neoplásica/genética , Neoplasias/genética , Comunicação Celular/genética , Comunicação Celular/imunologia , Humanos , Imunidade Inata/genética , Inflamação/imunologia , Inflamação/terapia , Metástase Neoplásica/imunologia , Metástase Neoplásica/terapia , Neoplasias/imunologia , Neoplasias/terapia , Microambiente Tumoral/genética , Microambiente Tumoral/imunologia
12.
PLoS One ; 14(5): e0216893, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31120919

RESUMO

CD4+ effector/memory T cells (Tem) represent a leading edge of the adaptive immune system responsible for protecting the body from infection, cancer, and other damaging processes. However, a subset of Tem cells with low expression of CD45Rb (RbLoTem) has been shown to suppress inflammation despite their effector surface phenotype and the lack of FoxP3 expression, the canonical transcription factor found in most regulatory T cells. In this report, we show that RbLoTem cells can suppress inflammation by influencing Treg behavior. Co-culturing activated RbLoTem and Tregs induced high expression of IL-10 in vitro, and conditioned media from RbLoTem cells induced IL-10 expression in FoxP3+ Tregs in vitro and in vivo, indicating that RbLoTem cells communicate with Tregs in a cell-contact independent fashion. Transcriptomic and multi-analyte Luminex data identified both IL-2 and IL-4 as potential mediators of RbLoTem-Treg communication, and antibody-mediated neutralization of either IL-4 or CD124 (IL-4Rα) prevented IL-10 induction in Tregs. Moreover, isolated Tregs cultured with recombinant IL-2 and IL-4 strongly induced IL-10 production. Using house dust mite (HDM)-induced airway inflammation as a model, we confirmed that the in vivo suppressive activity of RbLoTem cells was lost in IL-4-ablated RbLoTem cells. These data support a model in which RbLoTem cells communicate with Tregs using a combination of IL-2 and IL-4 to induce robust expression of IL-10 and suppression of inflammation.


Assuntos
Asma/imunologia , Comunicação Celular/imunologia , Interleucina-10/imunologia , Interleucina-4/imunologia , Antígenos Comuns de Leucócito/imunologia , Modelos Imunológicos , Linfócitos T Reguladores/imunologia , Animais , Asma/genética , Asma/patologia , Comunicação Celular/genética , Modelos Animais de Doenças , Inflamação/genética , Inflamação/imunologia , Inflamação/patologia , Interleucina-10/genética , Interleucina-2/genética , Interleucina-2/imunologia , Interleucina-4/genética , Antígenos Comuns de Leucócito/genética , Camundongos , Camundongos Knockout , Pyroglyphidae/imunologia , Linfócitos T Reguladores/patologia
13.
Glia ; 67(8): 1598-1619, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31033038

RESUMO

Diverse studies have suggested that cytoplasmic inclusions of misfolded α-synuclein in neuronal and glial cells are main pathological features of different α-synucleinopathies, including Parkinson's disease and dementia with Lewy bodies. Up to now, most studies have focused on the effects of α-synuclein on neurons, whereas the possible alterations of astrocyte functions and neuron-glia crosstalk have received minor attention. Recent evidence indicates that cellular signaling mediated by hemichannels and pannexons is critical for astroglial function and dysfunction. These channels constitute a diffusional route of communication between the cytosol and the extracellular space and during pathological scenarios they may lead to homeostatic disturbances linked to the pathogenesis and progression of different diseases. Here, we found that α-synuclein enhances the opening of connexin 43 (Cx43) hemichannels and pannexin-1 (Panx1) channels in mouse cortical astrocytes. This response was linked to the activation of cytokines, the p38 MAP kinase, the inducible nitric oxide synthase, cyclooxygenase 2, intracellular free Ca2+ concentration ([Ca2+ ]i ), and purinergic and glutamatergic signaling. Relevantly, the α-synuclein-induced opening of hemichannels and pannexons resulted in alterations in [Ca2+ ]i dynamics, nitric oxide (NO) production, gliotransmitter release, mitochondrial morphology, and astrocyte survival. We propose that α-synuclein-mediated opening of astroglial Cx43 hemichannels and Panx1 channels might constitute a novel mechanism involved in the pathogenesis and progression of α-synucleinopathies.


Assuntos
Astrócitos/patologia , Morte Celular/genética , Conexina 43/genética , Conexinas/genética , Proteínas do Tecido Nervoso/genética , alfa-Sinucleína/genética , Animais , Canais de Cálcio/genética , Canais de Cálcio/metabolismo , Comunicação Celular/genética , Células Cultivadas , Citocinas/metabolismo , Camundongos , Mitocôndrias/genética , Mitocôndrias/ultraestrutura , Neurotransmissores/metabolismo , Óxido Nítrico/biossíntese , RNA Interferente Pequeno/genética
14.
Mol Biol Rep ; 46(3): 3597-3606, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30989558

RESUMO

The integration of cell communication and the transfer of signals from stimuli via transcription to translation and further to activation of new protein is crucial for appropriate metabolism and function of living organisms. The overall elucidation and the examination of these complex processes require multistep laboratory approaches in order to obtain results which will not only detect particular stage but also indicate the mechanisms lying upon this process. Such results will be reliable because they will cover multidirectional methods and approaches. The analysis of currently available results already provided with the conclusion that often single omics approach does not correspond with other expected information and may bring misinterpretations. That is why the integration of several "omics" is useful for searching entire explanations and answers as well as appropriate interpretation of obtained complex results. The hypothesis was stated that "from transcriptomics can not be concluded to proteomics". This review focuses on the reasons for the integration of transcriptomic, proteomic and other-omics analysis. Moreover it also describes the examples of clinical meanings and mentions some methods used in these approaches.


Assuntos
Comunicação Celular/fisiologia , Biologia Computacional/métodos , Comunicação Celular/genética , Perfilação da Expressão Gênica/métodos , Humanos , Metabolômica/métodos , Proteômica/métodos
15.
Methods Mol Biol ; 1958: 403-436, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30945231

RESUMO

Membrane proteins are essential vessels for cell communication both with other cells and noncellular structures. They modulate environment responses and mediate a myriad of biological processes. Dimerization and multimerization processes have been shown to further increase the already high specificity of these processes. Due to their central role in various cell and organism functions, these multimers are often associated with health conditions, such as Alzheimer's disease (AD), Parkinson's disease (PD), and diabetes, among others.Understanding the membrane protein dimers' interface takes advantage of the specificity of the structure, for which we must pinpoint the most relevant interfacial residues, since they are extremely likely to be crucial for complex formation. Here, we describe step by step our own in silico protocol to characterize these residues, making use of known experimental structures. We detail the computational pipeline from data acquisition and pre-processing to feature extraction. A molecular dynamics simulation protocol to further study membrane dimer proteins and their interfaces is also illustrated.


Assuntos
Biologia Computacional/métodos , Proteínas de Membrana/química , Multimerização Proteica , Doença de Alzheimer/genética , Comunicação Celular/genética , Simulação por Computador , Diabetes Mellitus/genética , Humanos , Proteínas de Membrana/genética , Simulação de Dinâmica Molecular , Doença de Parkinson/genética , Ligação Proteica
16.
PLoS One ; 14(4): e0213069, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30947313

RESUMO

Extracellular vesicles (EVs) released by cells have a role in intercellular communication to regulate a wide range of biological processes. Two types of EVs can be recognized. Exosomes, which are released from multi-vesicular bodies upon fusion with the plasma membrane, and ectosomes, which directly bud from the plasma membrane. How cells regulate the quantity of EV release is largely unknown. One of the initiating events in vesicle biogenesis is the regulated transport of phospholipids from the exoplasmic to the cytosolic leaflet of biological membranes. This process is catalyzed by P4-ATPases. The role of these phospholipid transporters in intracellular vesicle transport has been established in lower eukaryotes and is slowly emerging in mammalian cells. In Caenorhabditis elegans (C. elegans), deficiency of the P4-ATPase member TAT-5 resulted in enhanced EV shedding, indicating a role in the regulation of EV release. In this study, we investigated whether the mammalian ortholog of TAT-5, ATP9A, has a similar function in mammalian cells. We show that knockdown of ATP9A expression in human hepatoma cells resulted in a significant increase in EV release that was independent of caspase-3 activation. Pharmacological blocking of exosome release in ATP9A knockdown cells did significantly reduce the total number of EVs. Our data support a role for ATP9A in the regulation of exosome release from human cells.


Assuntos
Adenosina Trifosfatases/genética , Exossomos/genética , Vesículas Extracelulares/genética , Proteínas de Membrana Transportadoras/genética , Animais , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/genética , Caspase 3/genética , Comunicação Celular/genética , Membrana Celular/genética , Micropartículas Derivadas de Células/genética , Endocitose/genética , Vesículas Extracelulares/metabolismo , Regulação da Expressão Gênica , Células Hep G2 , Humanos , Fosfolipídeos/metabolismo , Transporte Proteico/genética
17.
PLoS One ; 14(4): e0213422, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31017899

RESUMO

Epithelial wound healing requires the coordination of cells to migrate as a unit over the basement membrane after injury. To understand the process of this coordinated movement, it is critical to study the dynamics of cell-cell communication. We developed a method to characterize the injury-induced sustained Ca2+ mobilizations that travel between cells for periods of time up to several hours. These events of communication are concentrated along the wound edge and are reduced in cells further away from the wound. Our goal was to delineate the role and contribution of these sustained mobilizations and using MATLAB analyses, we determined the probability of cell-cell communication events in both in vitro models and ex vivo organ culture models. We demonstrated that the injury response was complex and represented the activation of a number of receptors. In addition, we found that pannexin channels mediated the cell-cell communication and motility. Furthermore, the sustained Ca2+ mobilizations are associated with changes in cell morphology and motility during wound healing. The results demonstrate that both purinoreceptors and pannexins regulate the sustained Ca2+ mobilization necessary for cell-cell communication in wound healing.


Assuntos
Cálcio/metabolismo , Comunicação Celular/genética , Córnea/metabolismo , Cicatrização/genética , Membrana Basal/metabolismo , Membrana Basal/ultraestrutura , Membrana Celular/metabolismo , Membrana Celular/patologia , Movimento Celular/genética , Córnea/patologia , Córnea/ultraestrutura , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Humanos , Microscopia Confocal , Técnicas de Cultura de Órgãos , Transdução de Sinais/genética
18.
Cancer Sci ; 110(6): 1947-1958, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31012516

RESUMO

MicroRNA is expected to be a novel therapeutic tool for tumors. Gap junctions facilitate the transfer of microRNA, which exerts biological effects on tumor cells. However, the length of microRNA that can pass through certain gap junctions composed of specific connexin remains unknown. To address this question, the present study investigated the permeability of gap junctions composed of various connexins, including connexin 43, connexin 32 or connexin 37, to microRNAs consisting of 18-27 nucleotides in glioma cells and cervical cancer cells. Results indicated that all of the microRNAs were able to be transferred from donor glioma cells to neighboring cells through the connexin 43 composed gap junction, but not the gap junctions composed of connexin 32 or connexin 37, in cervical cancer cells. Downregulation of the function of gap junctions comprising connexin 43 by pharmacological inhibition and shRNA significantly decreased the transfer of these microRNAs. In contrast, gap junction enhancers and overexpression of connexin 43 effectively increased these transfers. In glioma cells, cell proliferation was inhibited by microRNA-34a. Additionally, these effects of microRNA-34a were significantly enhanced by overexpression of connexin 43 in U251 cells, indicating that gap junctions play an important role in the antitumor effect of microRNA by transfer of microRNA to neighboring cells. Our data are the first to clarify the pattern of microRNA transmission through gap junctions and provide novel insights to show that antitumor microRNAs should be combined with connexin 43 or a connexin 43 enhancer, not connexin 32 or connexin 37, in order to improve the therapeutic effect.


Assuntos
Comunicação Celular/genética , Proliferação de Células/genética , Junções Comunicantes/metabolismo , MicroRNAs/genética , Linhagem Celular Tumoral , Permeabilidade da Membrana Celular/genética , Técnicas de Cocultura , Conexina 43/genética , Conexinas/genética , Glioma/genética , Glioma/patologia , Células HeLa , Humanos , Interferência de RNA
19.
Oncogene ; 38(26): 5227-5238, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30890754

RESUMO

Extracellular vesicles (EVs) can carry a wide array of RNAs in the tumor microenvironment, and are crucial for communication between tumor and surrounding stromal cells, including endothelial cells. Piwi-interacting RNAs (piRNAs) are important regulators implicated in the pathogenesis of multiple myeloma (MM). However, little is understood about the role of piRNA-823 in intercellular communication between MM and endothelial cells. In this study, we found that piRNA-823 mainly accumulated in EVs from peripheral blood of MM patients and EVs derived from MM cells (MM-derived-EVs). Increased piRNA-823 expression was associated with late stages and poor prognosis of MM. The MM-derived-EVs effectively transferred piRNA-823 to EA.hy926 endothelial cells. The piRNA-823 mimic and inhibitor were designed to upregulate or to suppress the endogenous function of piRNA-823. Transfection with piRNA-823 mimic or treatment with MM-derived-EVs significantly promoted the proliferation, tube formation, and invasion of EA.hy926 cells by enhancing the expression of VEGF, IL-6, and ICAM-1 and attenuating apoptosis. EA.hy926 cells transfected with piRNA-823 mimic or pre-treated with MM-derived-EVs promoted the growth of xenograft MM in mice. In contrast, the transfection with piRNA-823 inhibitor or treatment with EVs from piRNA-823 inhibitor-transfected-MM cells had diametrically opposite effects. Our findings demonstrated that piRNA-823 carried by MM-derived-EVs is essential for the re-education of ECs toward a unique environment amenable to the growth of MM cells by altering its biological characteristics. Our findings may pave the way for the development of new piRNA-mediated prognostic stratification and therapeutic strategies for MM.


Assuntos
Carcinogênese/genética , Comunicação Celular/genética , Células Endoteliais/fisiologia , Vesículas Extracelulares/fisiologia , Mieloma Múltiplo/patologia , RNA Interferente Pequeno/fisiologia , Microambiente Tumoral/fisiologia , Animais , Carcinogênese/efeitos dos fármacos , Células Cultivadas , Meios de Cultivo Condicionados/metabolismo , Meios de Cultivo Condicionados/farmacologia , Vesículas Extracelulares/genética , Vesículas Extracelulares/metabolismo , Vesículas Extracelulares/patologia , Feminino , Células Endoteliais da Veia Umbilical Humana , Humanos , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Mieloma Múltiplo/genética , Mieloma Múltiplo/metabolismo
20.
J Dermatol Sci ; 93(3): 159-167, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30904353

RESUMO

BACKGROUND: Pigmentation is controlled by complex mechanisms. Evidence suggests that miRNAs can regulate pigmentation. However, the mechanism has not been fully elucidated. Objective In this study, we revealed a novel mechanism that regulates pigmentation involving exosomes, miRNAs and the crosstalk between keratinocytes and melanocytes. METHODS: The expression and localization of exosome specific marker TSG101 in keratinocytes and melanocytes; Changes of melanin content in melanocytes after co-culture of exosome and melanocytes; Expression changes of target gene TYR and its related genes and inhibitory effect of miR-330-5p on pigmentation were studied by using various molecular biological techniques. RESULTS: In this experiment, we used miR-330-5p in keratinocytes to verify the effect of keratinocyte derived exosome on melanocyte pigmentation. First, we found that keratinocytes secrete exosomes carrying miR-330-5p; moreover, greater miR-330-5p expression was found in exosomes derived from keratinocytes that overexpressed miR-330-5p. Second, we found that exosomes derived from keratinocytes with overexpression of miR-330-5p caused a significant increase in miR-330-5p in melanocytes. Finally, exosomes derived from keratinocytes that overexpressed miR-330-5p induced a significant decrease in the production of melanin and expression of TYR in melanocytes. Meanwhile, we overexpressed miR-330-5p in melanocytes, which also proved the inhibitory effect of miR-330-5p on pigmentation. CONCLUSION: These findings suggest that keratinocytes crosstalk with melanocytes in the epidermal melanin unit via exosomal miRNAs. These studies reveal an important role of exosomes in melanocyte pigmentation, which opens a new pathway of melanogenesis.


Assuntos
Comunicação Celular/genética , Queratinócitos/metabolismo , Melanócitos/metabolismo , MicroRNAs/metabolismo , Pigmentação da Pele/genética , Animais , Técnicas de Cultura de Células , Linhagem Celular , Técnicas de Cocultura , Proteínas de Ligação a DNA/metabolismo , Complexos Endossomais de Distribuição Requeridos para Transporte/metabolismo , Exossomos/metabolismo , Queratinócitos/citologia , Melaninas/metabolismo , Camundongos , Monofenol Mono-Oxigenase/genética , Monofenol Mono-Oxigenase/metabolismo , Fatores de Transcrição/metabolismo
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