RESUMO
Lattice cells (LCs) in the developing Drosophila retina change shape before attaining final form. Previously, we showed that repeated contraction and expansion of apical cell contacts affect these dynamics. Here, we describe another factor, the assembly of a Rho1-dependent medioapical actomyosin ring formed by nodes linked by filaments that contract the apical cell area. Cell area contraction alternates with relaxation, generating pulsatile changes in cell area that exert force on neighboring LCs. Moreover, Rho1 signaling is sensitive to mechanical changes, becoming active when tension decreases and cells expand, while the negative regulator RhoGAP71E accumulates when tension increases and cells contract. This results in cycles of cell area contraction and relaxation that are reciprocally synchronized between adjacent LCs. Thus, mechanically sensitive Rho1 signaling controls pulsatile medioapical actomyosin contraction and coordinates cell behavior across the epithelium. Disrupting the kinetics of pulsing can lead to developmental errors, suggesting this process controls cell shape and tissue integrity during epithelial morphogenesis of the retina.
Assuntos
Actomiosina , Drosophila , Olho , Animais , Citoesqueleto de Actina/fisiologia , Actomiosina/fisiologia , Citocinese , Drosophila/embriologia , Morfogênese , Olho/embriologia , Proteínas rho de Ligação ao GTP/fisiologia , Proteínas de Drosophila/fisiologia , Retina/citologiaRESUMO
Miro1, a mitochondrial Rho GTPase1, is a kind of mitochondrial outer membrane protein involved in the regulation of mitochondrial anterograde transport and its subcellular distribution. Mitochondria influence reproductive processes of mammals in some aspects. Mitochondria are important for oocyte maturation, fertilization and embryonic development. The purpose of this study was to evaluate whether Miro1 regulates mouse oocyte maturation by altering mitochondrial homeostasis. We showed that Miro1 was expressed in mouse oocyte at different maturation stages. Miro1 mainly distributed in the cytoplasm and around the spindle during oocyte maturation. Small interference RNA-mediated Miro1 depletion caused significantly abnormal distribution of mitochondria and endoplasmic reticulum as well as mitochondrial dysfunction, resulting in severely impaired germinal vesicle breakdown (GVBD) of mouse oocytes. For those oocytes which went through GVBD in the Miro1-depleted group, part of them were inhibited in meiotic prophase I stage with abnormal chromosome arrangement and scattered spindle length. Our results suggest that Miro1 is essential for maintaining the maturation potential of mouse oocyte.
Assuntos
Meiose , Mitocôndrias , Oócitos , Proteínas rho de Ligação ao GTP , Animais , Feminino , Camundongos , Gravidez , Homeostase , Mitocôndrias/fisiologia , Oócitos/fisiologia , Oogênese , Proteínas rho de Ligação ao GTP/fisiologiaRESUMO
The extracellular contractile injection systems (eCISs) are encoded in the genomes of a large number of bacteria and archaea. We have previously characterized the overall structure of Photorhabdus Virulence Cassette (PVC), a typical member of the eCIS family. PVC resembles the contractile tail of bacteriophages and exerts its action by the contraction of outer sheath and injection of inner tube plus central spike. Nevertheless, the biological function of PVC effectors and the mechanism of effector translocation are still lacking. By combining cryo-electron microscopy and functional experiments, here we show that the PVC effectors Pdp1 (a new family of widespread dNTP pyrophosphatase effector in eCIS) and Pnf (a deamidase effector) are loaded inside the inner tube lumen in a "Peas in the Pod" mode. Moreover, we observe that Pdp1 and Pnf can be directly injected into J774A.1 murine macrophage and kill the target cells by disrupting the dNTP pools and actin cytoskeleton formation, respectively. Our results provide direct evidence of how PVC cargoes are loaded and delivered directly into mammalian macrophages.
Assuntos
Photorhabdus/patogenicidade , Microscopia Crioeletrônica , Células HEK293 , Células HeLa , Humanos , Mutagênese Sítio-Dirigida , Fases de Leitura Aberta , Photorhabdus/genética , Pirofosfatases/fisiologia , Virulência , Proteínas rho de Ligação ao GTP/fisiologiaRESUMO
Tunnelling nanotubes (TNTs) are an emerging route of long-range intercellular communication that mediate cell-to-cell exchange of cargo and organelles and contribute to maintaining cellular homeostasis by balancing diverse cellular stresses. Besides their role in intercellular communication, TNTs are implicated in several ways in health and disease. Transfer of pathogenic molecules or structures via TNTs can promote the progression of neurodegenerative diseases, cancer malignancy, and the spread of viral infection. Additionally, TNTs contribute to acquiring resistance to cancer therapy, probably via their ability to rescue cells by ameliorating various pathological stresses, such as oxidative stress, reactive oxygen species (ROS), mitochondrial dysfunction, and apoptotic stress. Moreover, mesenchymal stem cells play a crucial role in the rejuvenation of targeted cells with mitochondrial heteroplasmy and oxidative stress by transferring healthy mitochondria through TNTs. Recent research has focussed on uncovering the key regulatory molecules involved in the biogenesis of TNTs. However further work will be required to provide detailed understanding of TNT regulation. In this review, we discuss possible associations with Rho GTPases linked to oxidative stress and apoptotic signals in biogenesis pathways of TNTs and summarize how intercellular trafficking of cargo and organelles, including mitochondria, via TNTs plays a crucial role in disease progression and also in rejuvenation/therapy.
Assuntos
Comunicação Celular , Estresse Oxidativo , Proteínas rho de Ligação ao GTP/fisiologia , Humanos , Mitocôndrias/metabolismo , Neoplasias/metabolismo , Neoplasias/patologia , Doenças Neurodegenerativas/metabolismo , Doenças Neurodegenerativas/patologia , Organelas/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Viroses/metabolismo , Viroses/patologiaRESUMO
In the CNS, oligodendrocyte precursor cells differentiate into oligodendrocytes to wrap their plasma membranes around neuronal axons, generating mature neural networks with myelin sheaths according to spatial and temporal patterns. While myelination is known to be one of the most dynamic cell morphological changes, the overall intrinsic and extrinsic molecular cues controlling myelination remain to be fully clarified. Here, we describe the biphasic roles of Rnd2, an atypical branch of the Rho family GTPase, in oligodendrocyte myelination during development and after maturation in mice. Compared with littermate controls, oligodendrocyte-specific Rnd2 knockout mice exhibit decreased myelin thickness at the onset of myelination but increased myelin thickness in the later period. Larger proportions of Rho kinase and its substrate Mbs, the signaling unit that negatively regulates oligodendrocyte myelination, are phosphorylated at the onset of myelination, while their smaller proportions are phosphorylated in the later period. In addition, we confirm the biphasic role of Rnd2 through experiments with oligodendrocyte-specific Rnd2 transgenic mice. We conclude that Rnd2 positively regulates myelination in the early myelinating period and negatively regulates myelination in the later period. This unique modulator thus plays different roles depending on the myelination period.
Assuntos
Bainha de Mielina/metabolismo , Oligodendroglia/metabolismo , Proteínas rho de Ligação ao GTP/metabolismo , Animais , Diferenciação Celular/fisiologia , Células Cultivadas , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fibras Nervosas Mielinizadas/metabolismo , Neurônios/metabolismo , Organogênese , Transdução de Sinais , Proteínas rho de Ligação ao GTP/genética , Proteínas rho de Ligação ao GTP/fisiologiaRESUMO
Erythropoiesis is an intricate process starting in hematopoietic stem cells and leading to the daily production of 200 billion red blood cells (RBCs). Enucleation is a greatly complex and rate-limiting step during terminal maturation of mammalian RBC production involving expulsion of the nucleus from the orthochromatic erythroblasts, resulting in the formation of reticulocytes. The dynamic enucleation process involves many factors ranging from cytoskeletal proteins to transcription factors to microRNAs. Lack of optimum terminal erythroid maturation and enucleation has been an impediment to optimum RBC production ex vivo. Major efforts in the past two decades have exposed some of the mechanisms that govern the enucleation process. This review focuses in detail on mechanisms implicated in enucleation and discusses the future perspectives of this fascinating process.
Assuntos
Núcleo Celular , Eritroblastos/ultraestrutura , Eritrócitos/ultraestrutura , Eritropoese , Reticulócitos/ultraestrutura , Animais , Aves/sangue , Cálcio/fisiologia , Cromatina/ultraestrutura , Ensaio de Unidades Formadoras de Colônias , Biologia Computacional , Citocinas/fisiologia , Proteínas do Citoesqueleto/fisiologia , Proteínas de Ligação a DNA/fisiologia , Eritroblastos/citologia , Eritrócitos/citologia , Peptídeos e Proteínas de Sinalização Intercelular/fisiologia , Mamíferos/sangue , Camundongos , MicroRNAs/fisiologia , Proteínas Proto-Oncogênicas/fisiologia , Receptores dos Hormônios Tireóideos/fisiologia , Proteínas Repressoras/fisiologia , Reticulócitos/citologia , Fatores de Transcrição/fisiologia , Vesículas Transportadoras/fisiologia , Saco Vitelino/citologia , Proteínas rho de Ligação ao GTP/fisiologiaRESUMO
Small G-proteins of Rho family modulate the activity of several classes of ion channels, including K+ channels Kv1.2, Kir2.1, and ERG; Ca2+ channels; and epithelial Na+ channels. The present study was aimed to check the RhoA potential regulatory effects on Na+ current (INa) transferred by Na+ channel cardiac isoform NaV1.5 in heterologous expression system and in native rat cardiomyocytes. Whole-cell patch-clamp experiments showed that coexpression of NaV1.5 with the wild-type RhoA in CHO-K1 cell line caused 2.7-fold decrease of INa density with minimal influence on steady-state activation and inactivation. This effect was reproduced by the coexpression with a constitutively active RhoA, but not with a dominant negative RhoA. In isolated ventricular rat cardiomyocytes, a 5-h incubation with the RhoA activator narciclasine (5 × 10-6 M) reduced the maximal INa density by 38.8%. The RhoA-selective inhibitor rhosin (10-5 M) increased the maximal INa density by 25.3%. Experiments with sharp microelectrode recordings in isolated right ventricular wall preparations showed that 5 × 10-6 M narciclasine induced a significant reduction of action potential upstroke velocity after 2 h of incubation. Thus, RhoA might be considered as a potential negative regulator of sodium channels cardiac isoform NaV1.5.
Assuntos
Miócitos Cardíacos/fisiologia , Canal de Sódio Disparado por Voltagem NAV1.5/metabolismo , Sódio/metabolismo , Proteínas rho de Ligação ao GTP/fisiologia , Potenciais de Ação , Animais , Células CHO , Cricetulus , Masculino , Ratos , Ratos WistarRESUMO
Epileptogenesis is a potential process. Mossy fibre sprouting (MFS) and synaptic plasticity promote epileptogenesis. Overexpression of repulsive guidance molecule a (RGMa) prevents epileptogenesis by inhibiting MFS. However, other aspects underlying the RGMa regulatory process of epileptogenesis have not been elucidated. We studied whether RGMa could be modulated by microRNAs and regulated RhoA in epileptogenesis. Using microRNA databases, we selected four miRNAs as potential candidates. We further experimentally confirmed miR-20a-5p as a RGMa upstream regulator. Then, in vitro, by manipulating miR-20a-5p and RGMa, we investigated the regulatory relationship between miR-20a-5p, RGMa and RhoA, and the effects of this pathway on neuronal morphology. Finally, in the epilepsy animal model, we determined whether the miR-20a-5p-RGMa-RhoA pathway influenced MFS and synaptic plasticity and then modified epileptogenesis. Our results showed that miR-20a-5p regulated RGMa and that RGMa regulated RhoA in vitro. Furthermore, in primary hippocampal neurons, the miR-20a-5p-RGMa-RhoA pathway regulated axonal growth and neuronal branching; in the PTZ-induced epilepsy model, silencing miR-20a-5p prevented epileptogenesis through RGMa-RhoA-mediated synaptic plasticity but did not change MFS. Overall, we concluded that silencing miR-20a-5p inhibits axonal growth and neuronal branching and prevents epileptogenesis through RGMa-RhoA-mediated synaptic plasticity in the PTZ-induced epilepsy model, thereby providing a possible strategy to prevent epileptogenesis.
Assuntos
Proteínas Ligadas por GPI/fisiologia , Proteínas de Membrana/fisiologia , MicroRNAs/genética , Proteínas do Tecido Nervoso/fisiologia , Plasticidade Neuronal/fisiologia , Convulsões/prevenção & controle , Proteínas rho de Ligação ao GTP/fisiologia , Regiões 3' não Traduzidas , Animais , Axônios/ultraestrutura , Células Cultivadas , Convulsivantes/toxicidade , Dependovirus/genética , Modelos Animais de Doenças , Feminino , Proteínas Ligadas por GPI/biossíntese , Proteínas Ligadas por GPI/genética , Regulação da Expressão Gênica , Inativação Gênica , Vetores Genéticos , Hipocampo/citologia , Masculino , Proteínas de Membrana/biossíntese , Proteínas de Membrana/genética , MicroRNAs/antagonistas & inibidores , Proteínas do Tecido Nervoso/biossíntese , Proteínas do Tecido Nervoso/genética , Plasticidade Neuronal/genética , Neurônios/ultraestrutura , Pentilenotetrazol/toxicidade , RNA/metabolismo , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/farmacologia , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Convulsões/induzido quimicamente , Convulsões/genética , Convulsões/fisiopatologia , Transdução de Sinais , Proteínas rho de Ligação ao GTP/biossíntese , Proteínas rho de Ligação ao GTP/genéticaRESUMO
BACKGROUND The dishevelled associated activator of morphogenesis (DAAM) family, consisting of DAAM1 and DAAM2, is an important component of the Wnt signal pathway. Previous studies have suggested that DAAM2 reduces Von Hippel-Lindau (VHL) expression by promoting its ubiquitination, but the correlation between DAAM and HIF-1alpha in hepatocellular carcinoma (HCC) has not been studied. MATERIAL AND METHODS In our study, expression of DAAM1 and DAAM2 in HCCs and tumor-adjacent liver tissues was assessed with qRT-PCR and immunohistochemistry. Correlations between DAAM1/2 and the clinicopathologic variables were evaluated with the Chi-square test. With univariate and multivariate analysis, we further evaluated the prognostic significance of DAAM1 and DAAM2. Using in vitro experiments, we assessed the functions of DAAM1 and DAAM2 in invasion and proliferation in different HCC cell lines and investigated their underlying mechanisms. RESULTS DAAM1 and 2 overexpression were 18.8% and 48.7%, respectively, of the whole cohort. mRNAs of DAAM2 in HCCs were substantially higher than mRNAs in liver tissues, while DAAM1 mRNA had no marked difference. High DAAM2 expression was notably associated with advanced T stage (P=0.032), TNM stage (P=0.032), and overall survival (OS) rate (P=0.004). DAAM 2 knockdown promoted VHL accumulation and subsequent HIF-1alpha down-regulation in HCC cells. In HCC specimens, DAAM2 expression was also negatively correlated with VHL and positively associated with HIF-1alpha. Moreover, HIF-1alpha was required in DAAM2-induced invasion of HCC cells. CONCLUSIONS DAAM2, rather than DAAM1, was able to predict prognosis of HCC. DAAM2 decreased VHL expression and consequently upregulated HIF-1alpha, eventually facilitating invasion of HCC.
Assuntos
Carcinoma Hepatocelular/patologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Neoplasias Hepáticas/patologia , Proteínas dos Microfilamentos/fisiologia , Invasividade Neoplásica/fisiopatologia , Regulação para Cima , Proteínas rho de Ligação ao GTP/fisiologia , Carcinoma Hepatocelular/metabolismo , Linhagem Celular Tumoral , Feminino , Técnicas de Silenciamento de Genes , Humanos , Neoplasias Hepáticas/metabolismo , Masculino , Proteínas dos Microfilamentos/genética , Pessoa de Meia-Idade , Prognóstico , RNA Mensageiro/genética , Proteína Supressora de Tumor Von Hippel-Lindau/metabolismo , Proteínas rho de Ligação ao GTP/genéticaRESUMO
The cell wall is essential for cell viability and pathogenesis of fungi. It was previously shown that the exocytosis landmark Sec3 is an effector of the cell wall integrity (CWI) master regulator Rho1 GTPase. However, disruption of the interaction between Sec3 and Rho1 did not inhibit exocytic secretion and cell growth. The physiological role of Sec3 in fungi is unclear. We have examined the growth, cell wall sensitivity, exocyst localization, and exocytic secretion of Sec3-binding deficient rho1 mutants and Rho1-binding deficient sec3 mutants. We found that the Sec3 N-terminal deletion mutant was defective in cell wall integrity. The cells harboring binding mutation between Rho1 and Sec3 N-terminus were sensitive to cell wall antagonists. We also found that the polarized localization of exocyst subunits was disrupted in these mutants. Our study demonstrates that the N-terminus of Sec3 mediates cell wall integrity in yeast. Pathogenic fungi may use similar regulatory mechanisms because components of the exocytic signaling pathways are conserved.
Assuntos
Parede Celular/fisiologia , Proteínas de Saccharomyces cerevisiae/fisiologia , Saccharomyces cerevisiae/metabolismo , Sítios de Ligação/genética , Transporte Biológico/genética , Exocitose/genética , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/ultraestrutura , Proteínas de Saccharomyces cerevisiae/química , Proteínas rho de Ligação ao GTP/fisiologiaRESUMO
In cell migration, polarization is the process by which a stationary cell breaks symmetry and initiates motion. Although a lot is known about the mechanisms involved in cell polarization, the role played by myosin contraction remains unclear. In addition, cell polarization by mechanical impact has received little attention. Here, we study the influence of myosin activity on cell polarization and the initiation of motion induced by mechanical cues using a computational model for keratocytes. The model accounts for cell deformation, the dynamics of myosin and the signaling protein RhoA (a member of the Rho GTPases family), as well as the forces acting on the actomyosin network. Our results show that the attainment of a steady polarized state depends on the strength of myosin down- or up-regulation and that myosin upregulation favors cell polarization. Our results also confirm the existence of a threshold level for cell polarization, which is determined by the level of polarization of the Rho GTPases at the time the external stimuli vanish. In all, this paper shows that capturing the interactions between the signaling proteins (Rho GTPases for keratocytes) and the compounds of the motile machinery in a moving cell is crucial to study cell polarization.
Assuntos
Polaridade Celular , Queratinócitos/fisiologia , Modelos Biológicos , Miosinas/fisiologia , Movimento Celular , Fenômenos Físicos , Estimulação Física , Proteínas rho de Ligação ao GTP/fisiologiaRESUMO
Only a small number of genes are bona fide oncogenes and tumor suppressors such as Ras, Myc, ß-catenin, p53, and APC. However, targeting these cancer drivers frequently fail to demonstrate sustained cancer remission. Tumor heterogeneity and evolution contribute to cancer resistance and pose challenges for cancer therapy due to differential genomic rearrangement and expression driving distinct tumor responses to treatments. Here we report that intratumor heterogeneity of Wnt/ß-catenin modulator δ-catenin controls individual cell behavior to promote cancer. The differential intratumor subcellular localization of δ-catenin mirrors its compartmentalization in prostate cancer xenograft cultures as result of mutation-rendered δ-catenin truncations. Wild-type and δ-catenin mutants displayed distinct protein interactomes that highlight rewiring of signal networks. Localization specific δ-catenin mutants influenced p120ctn-dependent Rho GTPase phosphorylation and shifted cells towards differential bFGF-responsive growth and motility, a known signal to bypass androgen receptor dependence. Mutant δ-catenin promoted Myc-induced prostate tumorigenesis while increasing bFGF-p38 MAP kinase signaling, ß-catenin-HIF-1α expression, and the nuclear size. Therefore, intratumor δ-catenin heterogeneity originated from genetic remodeling promotes prostate cancer expansion towards androgen independent signaling, supporting a neomorphism model paradigm for targeting tumor progression.
Assuntos
Adenocarcinoma/patologia , Cateninas/fisiologia , Proteínas de Neoplasias/fisiologia , Neoplasias da Próstata/patologia , Transporte Ativo do Núcleo Celular , Adenocarcinoma/genética , Animais , Cateninas/genética , Linhagem Celular Tumoral , Núcleo Celular/ultraestrutura , DNA de Neoplasias/genética , Transição Epitelial-Mesenquimal/genética , Éxons/genética , Fator 2 de Crescimento de Fibroblastos/farmacologia , Genes myc , Xenoenxertos , Humanos , Subunidade alfa do Fator 1 Induzível por Hipóxia/fisiologia , Metástase Linfática/genética , Sistema de Sinalização das MAP Quinases , Masculino , Camundongos , Camundongos Transgênicos , Mutação , Proteínas de Neoplasias/genética , Neoplasias da Próstata/genética , Análise de Sequência de DNA , Frações Subcelulares/química , beta Catenina/fisiologia , Proteínas rho de Ligação ao GTP/fisiologia , delta CateninaRESUMO
Collective cell migration plays crucial roles in tissue remodeling, wound healing, and cancer cell invasion. However, its underlying mechanism remains unknown. Previously, we showed that the RhoA-targeting guanine nucleotide exchange factor Solo (ARHGEF40) is required for tensile force-induced RhoA activation and proper organization of keratin-8/keratin-18 (K8/K18) networks. Here, we demonstrate that Solo knockdown significantly increases the rate at which Madin-Darby canine kidney cells collectively migrate on collagen gels. However, it has no apparent effect on the migratory speed of solitary cultured cells. Therefore, Solo decelerates collective cell migration. Moreover, Solo localized to the anteroposterior regions of cell-cell contact sites in collectively migrating cells and was required for the local accumulation of K8/K18 filaments in the forward areas of the cells. Partial Rho-associated protein kinase (ROCK) inhibition or K18 or plakoglobin knockdown also increased collective cell migration velocity. These results suggest that Solo acts as a brake for collective cell migration by generating pullback force at cell-cell contact sites via the RhoA-ROCK pathway. It may also promote the formation of desmosomal cell-cell junctions related to K8/K18 filaments and plakoglobin.
Assuntos
Movimento Celular/fisiologia , Transdução de Sinais/fisiologia , Proteínas rho de Ligação ao GTP/fisiologia , Quinases Associadas a rho/fisiologia , Amidas/farmacologia , Animais , Polaridade Celular , Colágeno , Citoesqueleto/fisiologia , Desmossomos/fisiologia , Cães , Géis , Técnicas de Silenciamento de Genes , Queratina-18/antagonistas & inibidores , Queratina-18/genética , Queratina-18/fisiologia , Queratina-8/antagonistas & inibidores , Queratina-8/genética , Queratina-8/fisiologia , Células Madin Darby de Rim Canino , Piridinas/farmacologia , Interferência de RNA , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/farmacologia , Estresse Mecânico , Imagem com Lapso de Tempo , gama Catenina/antagonistas & inibidores , gama Catenina/genética , gama Catenina/fisiologia , Proteínas rac1 de Ligação ao GTP/fisiologia , Proteínas rho de Ligação ao GTP/antagonistas & inibidores , Proteína rhoA de Ligação ao GTP/fisiologiaRESUMO
PURPOSE OF REVIEW: This review provides an up-to-date understanding of how peroxisome proliferator activated receptor γ (PPARγ) exerts its cardioprotective effect in the vasculature through its activation of novel PPARγ target genes in endothelium and vascular smooth muscle. RECENT FINDINGS: In vascular endothelial cells, PPARγ plays a protective role by increasing nitric oxide bioavailability and preventing oxidative stress. RBP7 is a PPARγ target gene enriched in vascular endothelial cells, which is likely to form a positive feedback loop with PPARγ. In vascular smooth muscle cells, PPARγ antagonizes the renin-angiotensin system, maintains vascular integrity, suppresses vasoconstriction, and promotes vasodilation through distinct pathways. Rho-related BTB domain containing protein 1 (RhoBTB1) is a novel PPARγ gene target in vascular smooth muscle cells that mediates the protective effect of PPARγ by serving as a substrate adaptor between the Cullin-3 RING ubiquitin ligase and phosphodiesterase 5, thus restraining its activity through ubiquitination and proteasomal degradation. SUMMARY: In the vasculature, PPARγ exerts its cardioprotective effect through its transcriptional activity in endothelium and vascular smooth muscle. From the understanding of PPARγ's transcription targets in those pathways, novel hypertension therapy target(s) will emerge.
Assuntos
Hipertensão/etiologia , PPAR gama/fisiologia , Proteínas rho de Ligação ao GTP/fisiologia , Animais , Células Endoteliais/fisiologia , Humanos , Músculo Liso Vascular/citologia , Miócitos de Músculo Liso/fisiologiaRESUMO
As membrane-associated master regulators of cytoskeletal remodeling, Rho GTPases coordinate a wide range of biological processes such as cell adhesion, motility, and polarity. In the last years, Rho GTPases have also been recognized to control intracellular membrane sorting and trafficking steps directly; however, how Rho GTPase signaling is regulated at endomembranes is still poorly understood. In this review, we will specifically address the local Rho GTPase pools coordinating intracellular membrane trafficking with a focus on the endo- and exocytic pathways. We will further highlight the spatiotemporal molecular regulation of Rho signaling at endomembrane sites through Rho regulatory proteins, the GEFs and GAPs. Finally, we will discuss the contribution of dysregulated Rho signaling emanating from endomembranes to the development and progression of cancer.
Assuntos
Membranas Intracelulares/metabolismo , Proteínas rho de Ligação ao GTP/metabolismo , Proteínas rho de Ligação ao GTP/fisiologia , Adesão Celular , Movimento Celular , Polaridade Celular , Citoesqueleto/metabolismo , Endocitose/fisiologia , Exocitose/fisiologia , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Humanos , Transporte Proteico/fisiologia , Transdução de SinaisRESUMO
BACKGROUND: Adipose tissue plays a crucial role in diet- and obesity-related insulin resistance, with implications for several metabolic diseases. Identification of novel target genes and mechanisms that regulate adipocyte function could lead to improved treatment strategies. RND3 (RhoE/Rho8), a Rho-related GTP-binding protein that inhibits Rho kinase (ROCK) signaling, has been linked to diverse diseases such as apoptotic cardiomyopathy, heart failure, cancer and type 2 diabetes, in part by regulating cytoskeleton dynamics and insulin-mediated glucose uptake. RESULTS: We here investigated the expression of RND3 in adipose tissue in human obesity, and discovered a role for RND3 in regulating adipocyte metabolism. In cross-sectional and prospective studies, we observed 5-fold increased adipocyte levels of RND3 mRNA in obesity, reduced levels after surgery-induced weight loss, and positive correlations of RND3 mRNA with adipocyte size and surrogate measures of insulin resistance (HOMA2-IR and circulating triglyceride/high-density lipoprotein cholesterol (TAG/HDL-C) ratio). By screening for RND3-dependent gene expression following siRNA-mediated RND3 knockdown in differentiating human adipocytes, we found downregulation of inflammatory genes and upregulation of genes related to adipocyte ipolysis and insulin signaling. Treatment of adipocytes with tumor necrosis factor alpha (TNFα), lipopolysaccharide (LPS), hypoxia or cAMP analogs increased RND3 mRNA levels 1.5-2-fold. Functional assays in primary human adipocytes confirmed that RND3 knockdown reduces cAMP- and isoproterenol-induced lipolysis, which were mimicked by treating cells with ROCK inhibitor. This effect could partly be explained by reduced protein expression of adipose triglyceride lipase (ATGL) and phosphorylated hormone-sensitive lipase (HSL). CONCLUSION: We here uncovered a novel differential expression of adipose RND3 in obesity and insulin resistance, which may at least partly depend on a causal effect of RND3 on adipocyte lipolysis.
Assuntos
Adipócitos/metabolismo , Lipólise/efeitos dos fármacos , Proteínas rho de Ligação ao GTP/fisiologia , Animais , Células Cultivadas , Estudos Transversais , Regulação da Expressão Gênica , Humanos , Resistência à Insulina , Obesidade/metabolismo , Estudos Prospectivos , RNA Mensageiro/metabolismo , Proteínas rho de Ligação ao GTP/genética , Proteínas rho de Ligação ao GTP/metabolismoRESUMO
Podocytes, or glomerular epithelial cells, form the final layer in the glomerular capillary wall of the kidney. Along with the glomerular basement membrane and glomerular endothelial cells, they make up the glomerular filtration barrier which allows the passage of water and small molecules and, in healthy individuals, prevents the passage of albumin and other key proteins. The podocyte is a specialised and terminally differentiated cell with a specific cell morphology that is largely dependent on a highly dynamic underlying cytoskeletal network and that is essential for maintaining glomerular function and integrity in healthy kidneys. The RhoGTPases (RhoA, Rac1 and Cdc42), which act as molecular switches that regulate actin dynamics, are known to play a crucial role in maintaining the cytoskeletal and molecular integrity of the podocyte foot processes in a dynamic manner. Recently, novel protein interaction networks that regulate the RhoGTPases in the podocyte and that are altered by disease have been discovered. This review will discuss these networks and their potential as novel therapeutic targets in nephrotic syndrome. It will also discuss the evidence that they are direct targets for (a) steroids, the first-line agents for the treatment of nephrotic syndrome, and (b) certain kinase inhibitors used in cancer treatment, leading to nephrotoxicity.
Assuntos
Nefropatias , Síndrome Nefrótica , Podócitos , Proteínas rho de Ligação ao GTP , Citoesqueleto , Células Endoteliais , Humanos , Síndrome Nefrótica/tratamento farmacológico , Proteínas rho de Ligação ao GTP/fisiologia , Proteínas rho de Ligação ao GTP/uso terapêuticoRESUMO
INTRODUCTION: Erectile dysfunction (ED) is frequently encountered in patients with arterial hypertension and there is a recent functional correlation between the expression of thermoreceptor channels TRPM8 (melastatin 8) and alterations in blood pressure in hypertension. The aim of this study was to investigate the function of cold-sensing TRPM8 channel in internal pudendal artery (IPA) in both normotensive and hypertensive rats. METHODS: We performed experiments integrating physiological, pharmacological, biochemical and cellular techniques. RESULTS: TRPM8 channels are expressed in the IPA and in vascular smooth muscle cells from IPA. In addition, TRPM8 activation, by both a cooling compound icilin (82.1⯱â¯3.0%, nâ¯=â¯6) and cold temperature [thermal stimulus, basal tone (25⯰C, 41.2⯱â¯3.4%, nâ¯=â¯5) or pre-contracted tone induced by phenylephrine (25⯰C, 87.0⯱â¯3.6%, nâ¯=â¯7)], induced relaxation in IPA. Furthermore, the results showed that the concentration-response curve to icilin was significantly shifted to the right in different conditions, such as: the absence of the vascular endothelium, in the presence of L-NAME (10-4 M), or indomethacin (10-5 M) or by a combination of charybdotoxin (10-7 M) and apamin (5â¯×â¯10-6 M), and Y27632 (10-6 M). Interestingly, icilin-induced vasodilation was significantly higher in IPA from spontaneously hypertensive (SHR, E10-4M = 75.3⯱â¯1.7%) compared to wistar rats (E10-4M = 56.4⯱â¯2.6%), despite no changes in the TRPM8 expression in IPA between the strains, suggesting that the sensitivity of TRPM8 channels is higher in SHR. CONCLUSIONS: These data demonstrate for the first time, the expression and function of TRPM8 channels in the IPA involving, at least in part, endothelium-derived relaxing factors and ROCK inhibition. Overall, this channel could potentially be a new target for the treatment of hypertension associated-ED.
Assuntos
Artérias/fisiologia , Hipertensão/fisiopatologia , Canais de Cátion TRPM/fisiologia , Animais , Masculino , Músculo Liso Vascular/fisiologia , Miócitos de Músculo Liso/fisiologia , Ratos Endogâmicos SHR , Ratos Sprague-Dawley , Ratos Wistar , Vasodilatação , Proteínas rho de Ligação ao GTP/fisiologia , Quinases Associadas a rho/fisiologiaRESUMO
SB-706375 is a selective receptor antagonist of human urotensin-II (hU-II), which can block the aorta contraction induced by hU-II in rats. The effect of SB-706375 on myocardial ischaemia-reperfusion (I/R) injury is unclear. The major objective of this study was to investigate whether SB-706375 has a protective effect on myocardial I/R injury in rats and explore its possible mechanisms. Isolated hearts of Adult Sprague-Dawley were perfused in a Langendorff apparatus, and haemodynamic parameters, lactate dehydrogenase (LDH), creatine phosphokinase-MB (CK-MB), cardiac troponin I (cTnI), RhoA, and the protein expressions of U-II receptor (UTR), receptor-interacting protein 3 (RIP3), Rho-associated coiled-coil-containing protein kinase 1 (ROCK1) and Rho-associated coiled-coil-containing protein kinase 2 (ROCK2) were assessed. We found that SB-706375 (1 × 10-6 and 1 × 10-5 mol/L) significantly inhibited the changes of haemodynamic parameters and reduced LDH and CK-MB activities and also cTnI level in the coronary effluents in the heart subjected to myocardial I/R injury. Further experiments studies showed that SB-706375 obviously prevented myocardial I/R increased RhoA activity and UTR, RIP3, ROCK1, and ROCK2 protein expressions. ROCK inhibition abolished the improving effect of SB-706375 on myocardial I/R-induced haemodynamic change in the isolated perfused rat heart. These findings suggested that SB-706375 provides cardio-protection against I/R injury in isolated rats by blocking UTR-RhoA/ROCK-RIP3 pathway.
Assuntos
Traumatismo por Reperfusão Miocárdica/prevenção & controle , Miocárdio/patologia , Pirrolidinas/farmacologia , Proteína Serina-Treonina Quinases de Interação com Receptores/antagonistas & inibidores , Receptores Acoplados a Proteínas G/antagonistas & inibidores , Sulfonamidas/farmacologia , Proteínas rho de Ligação ao GTP/antagonistas & inibidores , Quinases Associadas a rho/antagonistas & inibidores , Animais , Feminino , Masculino , Necrose , Ratos , Ratos Sprague-Dawley , Proteína Serina-Treonina Quinases de Interação com Receptores/fisiologia , Receptores Acoplados a Proteínas G/análise , Transdução de Sinais/fisiologia , Proteínas rho de Ligação ao GTP/fisiologia , Quinases Associadas a rho/fisiologiaRESUMO
Stroke significantly affects white matter in the brain by impairing axon function, which results in clinical deficits. Axonal mitochondria are highly dynamic and are transported via microtubules in the anterograde or retrograde direction, depending upon axonal energy demands. Recently, we reported that mitochondrial division inhibitor 1 (Mdivi-1) promotes axon function recovery by preventing mitochondrial fission only when applied during ischemia. Application of Mdivi-1 after injury failed to protect axon function. Interestingly, L-NIO, which is a NOS3 inhibitor, confers post-ischemic protection to axon function by attenuating mitochondrial fission and preserving mitochondrial motility via conserving levels of the microtubular adaptor protein Miro-2. We propose that preventing mitochondrial fission protects axon function during injury, but that restoration of mitochondrial motility is more important to promote axon function recovery after injury. Thus, Miro-2 may be a therapeutic molecular target for recovery following a stroke.