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1.
Biol Res ; 55(1): 35, 2022 Nov 26.
Artículo en Inglés | MEDLINE | ID: mdl-36435789

RESUMEN

Extracellular vesicles (EVs) are naturally released membrane vesicles that act as carriers of proteins and RNAs for intercellular communication. With various biomolecules and specific ligands, EV has represented a novel form of information transfer, which possesses extremely outstanding efficiency and specificity compared to the classical signal transduction. In addition, EV has extended the concept of signal transduction to intercellular aspect by working as the collection of extracellular information. Therefore, the functions of EVs have been extensively characterized and EVs exhibit an exciting prospect for clinical applications. However, the biogenesis of EVs and, in particular, the regulation of this process by extracellular signals, which are essential to conduct further studies and support optimal utility, remain unclear. Here, we review the current understanding of the biogenesis of EVs, focus on the regulation of this process by extracellular signals and discuss their therapeutic value.


Asunto(s)
Vesículas Extracelulares , Vesículas Extracelulares/metabolismo , Comunicación Celular/fisiología , Transducción de Señal , Transporte Biológico , ARN/metabolismo
2.
Biofabrication ; 16(3)2024 Apr 03.
Artículo en Inglés | MEDLINE | ID: mdl-38471164

RESUMEN

Cells sense mechanical signals from the surrounding environment and transmit them to the nucleus through mechanotransduction to regulate cellular behavior. Microcontact printing, which utilizes elastomer stamps, is an effective method for simulating the cellular microenvironment and manipulating cell morphology. However, the conventional fabrication process of silicon masters and elastomer stamps requires complex procedures and specialized equipment, which restricts the widespread application of micropatterning in cell biology and hinders the investigation of the role of cell geometry in regulating cell behavior. In this study, we present an innovative method for convenient resin stamp microfabrication based on digital micromirror device planar lithography. Using this method, we generated a series of patterns ranging from millimeter to micrometer scales and validated their effectiveness in controlling adhesion at both collective and individual cell levels. Additionally, we investigated mechanotransduction and cell behavior on elongated micropatterned substrates. We then examined the effects of cell elongation on cytoskeleton organization, nuclear deformation, focal adhesion formation, traction force generation, nuclear mechanics, and the growth of HeLa cells. Our findings reveal a positive correlation between cell length and mechanotransduction. Interestingly, HeLa cells with moderate length exhibit the highest cell division and proliferation rates. These results highlight the regulatory role of cell elongation in mechanotransduction and its significant impact on cancer cell growth. Furthermore, our methodology for controlling cell adhesion holds the potential for addressing fundamental questions in both cell biology and biomedical engineering.


Asunto(s)
Elastómeros , Mecanotransducción Celular , Humanos , Células HeLa , Adhesión Celular/fisiología , División Celular
3.
Front Med ; 18(3): 430-445, 2024 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-38600350

RESUMEN

Crosstalk between cancer cells and the immune microenvironment is determinant for liver cancer progression. A tumor subpopulation called liver cancer stem cells (CSCs) significantly accounts for the initiation, metastasis, therapeutic resistance, and recurrence of liver cancer. Emerging evidence demonstrates that the interaction between liver CSCs and immune cells plays a crucial role in shaping an immunosuppressive microenvironment and determining immunotherapy responses. This review sheds light on the bidirectional crosstalk between liver CSCs and immune cells for liver cancer progression, as well as the underlying molecular mechanisms after presenting an overview of liver CSCs characteristic and their microenvironment. Finally, we discuss the potential application of liver CSCs-targeted immunotherapy for liver cancer treatment.


Asunto(s)
Progresión de la Enfermedad , Inmunoterapia , Neoplasias Hepáticas , Células Madre Neoplásicas , Microambiente Tumoral , Humanos , Microambiente Tumoral/inmunología , Células Madre Neoplásicas/inmunología , Neoplasias Hepáticas/inmunología , Neoplasias Hepáticas/patología , Comunicación Celular/inmunología
4.
Cancer Lett ; 596: 216961, 2024 Aug 01.
Artículo en Inglés | MEDLINE | ID: mdl-38823764

RESUMEN

Extracellular vesicles are essential for intercellular communication and are involved in tumor progression. Inhibiting the direct release of extracellular vesicles seems to be an effective strategy in inhibiting tumor progression, but lacks of investigation. Here, we report a natural flavonoid compound, apigenin, could significantly inhibit the growth of hepatocellular carcinoma by preventing microvesicle secretion. Mechanistically, apigenin primarily targets the guanine nucleotide exchange factor ARHGEF1, inhibiting the activity of small G protein Cdc42, which is essential in regulating the release of microvesicles from tumor cells. In turn, this inhibits tumor angiogenesis related to VEGF90K transported on microvesicles, ultimately impeding tumor progression. Collectively, these findings highlight the therapeutic potential of apigenin and shed light on its anticancer mechanisms through inhibiting microvesicle biogenesis, providing a solid foundation for the refinement and practical application of apigenin.


Asunto(s)
Apigenina , Carcinoma Hepatocelular , Micropartículas Derivadas de Células , Neoplasias Hepáticas , Neovascularización Patológica , Factores de Intercambio de Guanina Nucleótido Rho , Humanos , Neovascularización Patológica/tratamiento farmacológico , Neovascularización Patológica/metabolismo , Animales , Apigenina/farmacología , Neoplasias Hepáticas/tratamiento farmacológico , Neoplasias Hepáticas/patología , Neoplasias Hepáticas/metabolismo , Factores de Intercambio de Guanina Nucleótido Rho/metabolismo , Factores de Intercambio de Guanina Nucleótido Rho/genética , Micropartículas Derivadas de Células/metabolismo , Micropartículas Derivadas de Células/efectos de los fármacos , Carcinoma Hepatocelular/tratamiento farmacológico , Carcinoma Hepatocelular/patología , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/irrigación sanguínea , Ratones , Línea Celular Tumoral , Proteína de Unión al GTP cdc42/metabolismo , Proliferación Celular/efectos de los fármacos , Ensayos Antitumor por Modelo de Xenoinjerto , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Células Endoteliales de la Vena Umbilical Humana/efectos de los fármacos , Células Hep G2 , Ratones Desnudos , Angiogénesis
5.
Nat Cell Biol ; 8(9): 945-56, 2006 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-16892055

RESUMEN

Cool-1 (cloned-out of library 1) has a key role in regulating epidermal growth factor receptor (EGFR) degradation. Here, we show that Cool-1 performs this function by functioning as both an upstream activator and downstream target for Cdc42. EGF-dependent phosphorylation of Cool-1 enables it to act as a nucleotide exchange factor for Cdc42 and to form a complex with the E3 ligase Cbl, thus regulating Cbl-catalysed EGFR degradation. The EGF-dependent phosphorylation is normally transient; however, Cool-1 phosphorylation is sustained in cells expressing v-Src and is essential for cellular transformation, as well as for v-Src-induced tumour formation in mice. These findings demonstrate that the regulated phosphorylation of Cool-1 is necessary to maintain the balance between normal signalling by EGFR and Src versus aberrant growth and transformation.


Asunto(s)
Proteínas de Ciclo Celular/fisiología , Proliferación Celular , Transformación Celular Neoplásica , Receptores ErbB/fisiología , Factores de Intercambio de Guanina Nucleótido/fisiología , Proteína Oncogénica pp60(v-src)/fisiología , Proteína de Unión al GTP cdc42/fisiología , Animales , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Endocitosis , Factores de Intercambio de Guanina Nucleótido/genética , Factores de Intercambio de Guanina Nucleótido/metabolismo , Masculino , Ratones , Ratones Desnudos , Células 3T3 NIH , Trasplante de Neoplasias , Neoplasias Experimentales/patología , Proteína Oncogénica pp60(v-src)/biosíntesis , Fosforilación , Fosfotirosina/metabolismo , Unión Proteica , Proteínas Proto-Oncogénicas c-cbl/metabolismo , Interferencia de ARN , Factores de Intercambio de Guanina Nucleótido Rho , Transducción de Señal , Trasplante Heterólogo
6.
Cancer Lett ; 571: 216348, 2023 09 01.
Artículo en Inglés | MEDLINE | ID: mdl-37567461

RESUMEN

Copper is a necessary cofactor vital for maintaining biological functions, as well as participating in the development of cancer. A plethora of studies have demonstrated that copper is a double-edged sword, presenting both benefits and detriments to tumors. The liver is a metabolically active organ, and an imbalance of copper homeostasis can result in deleterious consequences to the liver. Hepatocellular carcinoma (HCC), the most common primary liver cancer, is a highly aggressive malignancy with limited viable therapeutic options. As research advances, the focus has shifted towards the relationships between copper and HCC. Innovatively, cuproplasia and cuproptosis have been proposed to depict copper-related cellular growth and death, providing new insights for HCC treatment. By summarizing the constantly elucidated molecular connections, this review discusses the mechanisms of copper in the pathogenesis, progression, and potential therapeutics of HCC. Additionally, we aim to tentatively provide a theoretical foundation and gospel for HCC patients.


Asunto(s)
Carcinoma Hepatocelular , Neoplasias Hepáticas , Humanos , Carcinoma Hepatocelular/tratamiento farmacológico , Carcinoma Hepatocelular/genética , Cobre , Neoplasias Hepáticas/tratamiento farmacológico , Neoplasias Hepáticas/genética , Proliferación Celular
7.
Sci Adv ; 9(49): eadi2465, 2023 12 08.
Artículo en Inglés | MEDLINE | ID: mdl-38055816

RESUMEN

Adenosine 5'-triphosphate citrate lyase (ACLY) is a cytosolic enzyme that converts citrate into acetyl-coenzyme A for fatty acid and cholesterol biosynthesis. ACLY is up-regulated or activated in many cancers, and targeting ACLY by inhibitors holds promise as potential cancer therapy. However, the role of ACLY in cancer immunity regulation remains poorly understood. Here, we show that ACLY inhibition up-regulates PD-L1 immune checkpoint expression in cancer cells and induces T cell dysfunction to drive immunosuppression and compromise its antitumor effect in immunocompetent mice. Mechanistically, ACLY inhibition causes polyunsaturated fatty acid (PUFA) peroxidation and mitochondrial damage, which triggers mitochondrial DNA leakage to activate the cGAS-STING innate immune pathway. Pharmacological and genetic inhibition of ACLY overcomes cancer resistance to anti-PD-L1 therapy in a cGAS-dependent manner. Furthermore, dietary PUFA supplementation mirrors the enhanced efficacy of PD-L1 blockade by ACLY inhibition. These findings reveal an immunomodulatory role of ACLY and provide combinatorial strategies to overcome immunotherapy resistance in tumors.


Asunto(s)
Antígeno B7-H1 , Neoplasias , Ratones , Animales , Neoplasias/tratamiento farmacológico , Neoplasias/metabolismo , Ácidos Grasos Insaturados , Nucleotidiltransferasas , Inmunoterapia
8.
Heliyon ; 8(11): e11503, 2022 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-36411886

RESUMEN

Metformin is a drug that has been applied in clinical use for many years for the treatment of type 2 diabetes mellitus (T2DM). It achieves its function through multiple targets and modulation of multiple signaling pathways. To date, the mechanism of the action of metformin is still not fully understood. Along with glycemic control, metformin has shown good inhibitory effects on the development of many tumors. Here, we elucidated that plasma exosomal microRNA-122-5p (miR-122) is closely related to the mechanism of metformin. MiR-122 regulates glycogen-glucose metabolism in hepatocytes or hepatocellular carcinoma cells (HCC) by inhibiting the phosphorylation of AMPK. Since miR-122 and metformin regulate glucose metabolism homeostasis through similar mechanisms, miR-122 can antagonize the effects of metformin. MiR-122 expression increases the sensitivity of hepatocytes or HCC to metformin. Conversely, decreased expression of miR-122 results in hepatocyte insensitivity to metformin. Therefore, significantly elevated levels of miR-122 in plasma exosomes of hepatocellular carcinoma patients could enhance their sensitivity to metformin. The results of the present study revealed a key regulatory role of plasma exosomal miR-122 on the molecular mechanism of metformin. The regulation of key molecules of related signaling pathways by miR-122 may lead to similar glycemic lowering and tumor suppression therapeutic effects as metformin. This provides new ideas for the development of new therapeutic strategies for hepatocellular carcinoma based on the mechanism of miR-122 and metformin.

9.
J Biol Chem ; 285(24): 18806-16, 2010 Jun 11.
Artículo en Inglés | MEDLINE | ID: mdl-20375009

RESUMEN

Previously we showed that Cool-1 (Cloned out of library-1)/beta-Pix (Pak-interactive exchange factor) is phosphorylated at a specific tyrosine residue (Tyr-442) in a Src-dependent manner and serves as a dual function guanine nucleotide exchange factor (GEF)/signaling-effector for Cdc42 that is essential for transformation by Src. Here, we show that knocking-down Cool-1 or overexpressing a Cool-1 mutant that contains substitutions within its Dbl homology domain and is defective for GEF activity, inhibits Src-promoted cell migration. Similarly, the expression of a Cool-1 mutant containing a tyrosine to phenylalanine substitution at position 442, making it incapable of being phosphorylated in response to serum, epidermal growth factor (EGF), or Src, also causes a significant inhibition of the migration and invasive activity of cells expressing oncogenic Src. We further demonstrate that the phosphorylation of Cool-1 at Tyr-442 weakens its ability to bind to one of its primary interaction-partners, Cat-1 (Cool-associated tyrosine phosphosubstrate-1)/Git-1 (G protein-coupled receptor kinase-interactor-1), thus making Cat more accessible for binding to paxillin. This enables cells to alternate between states where they contain large numbers of focal complexes (i.e. conditions favoring Cool-1-Cat interactions) versus reduced numbers of focal complexes (conditions favoring Cat-paxillin interactions). Overall, these findings show that the phosphorylation-dephosphorylation cycle of Cool-1 at Tyr-442 can serve as a key regulatory signal for focal complex assembly-disassembly, and consequently, for the migration and invasive activity of Src-transformed cells.


Asunto(s)
Factores de Intercambio de Guanina Nucleótido/metabolismo , Animales , Membrana Celular/metabolismo , Movimiento Celular , Glutatión Transferasa/metabolismo , Ratones , Microscopía Fluorescente/métodos , Mutación , Células 3T3 NIH , Invasividad Neoplásica , Fosforilación , Estructura Terciaria de Proteína , Factores de Intercambio de Guanina Nucleótido Rho , Tirosina/química , Proteína de Unión al GTP cdc42/metabolismo
10.
J Biol Chem ; 285(8): 5815-26, 2010 Feb 19.
Artículo en Inglés | MEDLINE | ID: mdl-20028979

RESUMEN

Although the small GTPase Ran is best known for its roles in nucleocytoplasmic transport, mitotic spindle assembly, and nuclear envelope formation, recent studies have demonstrated the overexpression of Ran in multiple tumor types and that its expression is correlated with a poor patient prognosis, providing evidence for the importance of this GTPase in cell growth regulation. Here we show that Ran is subject to growth factor regulation by demonstrating that it is activated in a serum-dependent manner in human breast cancer cells and, in particular, in response to heregulin, a growth factor that activates the Neu/ErbB2 tyrosine kinase. The heregulin-dependent activation of Ran requires mTOR (mammalian target of rapamycin) and stimulates the capped RNA binding capability of the cap-binding complex in the nucleus, thus influencing gene expression at the level of mRNA processing. We further demonstrate that the excessive activation of Ran has important consequences for cell growth by showing that a novel, activated Ran mutant is sufficient to transform NIH-3T3 cells in an mTOR- and epidermal growth factor receptor-dependent manner and that Ran-transformed cells form tumors in mice.


Asunto(s)
Neoplasias de la Mama/enzimología , Núcleo Celular/enzimología , Transformación Celular Neoplásica/metabolismo , Neurregulina-1/metabolismo , Proteína de Unión al GTP ran/metabolismo , Animales , Neoplasias de la Mama/genética , Línea Celular Tumoral , Núcleo Celular/genética , Transformación Celular Neoplásica/genética , Activación Enzimática/efectos de los fármacos , Femenino , Humanos , Péptidos y Proteínas de Señalización Intracelular/genética , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Ratones , Mutación , Células 3T3 NIH , Neurregulina-1/farmacología , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas de Unión a Caperuzas de ARN/genética , Proteínas de Unión a Caperuzas de ARN/metabolismo , Procesamiento Postranscripcional del ARN/efectos de los fármacos , Procesamiento Postranscripcional del ARN/genética , ARN Mensajero/biosíntesis , ARN Mensajero/genética , Receptor ErbB-2/genética , Receptor ErbB-2/metabolismo , Serina-Treonina Quinasas TOR , Proteína de Unión al GTP ran/genética
11.
Front Immunol ; 12: 625472, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33763069

RESUMEN

Neutrophils are the largest population of circulating leukocytes and the first responder against invading pathogens or other danger signals. Sophisticated machineries help them play critical roles in immunity and inflammation, including phagocytosis, superoxide production, cytokine and chemokine production, degranulation, and formation of neutrophil extracellular traps (NETs). After maturation and release from the bone marrow, neutrophils migrate to inflamed tissues in response to many stimuli. Increasing evidences indicate that neutrophils are critically involved in the pathogenesis of liver diseases, including liver cancer, thus making them promising target for the treatment of liver diseases. Here, we would like to provide the latest finding about the role of neutrophils in liver diseases and discuss the potentiality of neutrophils as target for liver diseases.


Asunto(s)
Hepatopatías/inmunología , Hígado/inmunología , Activación Neutrófila , Infiltración Neutrófila , Neutrófilos/inmunología , Animales , Enfermedad Crónica , Citocinas/metabolismo , Humanos , Hígado/metabolismo , Hígado/patología , Hepatopatías/metabolismo , Hepatopatías/patología , Neutrófilos/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal
12.
J Extracell Vesicles ; 10(3): e12051, 2021 01.
Artículo en Inglés | MEDLINE | ID: mdl-33473262

RESUMEN

Tumour-derived microvesicles (MVs) serve as critical mediators of cell-to-cell communication in the tumour microenvironment. So far, the underlying mechanisms of MV biogenesis, especially how key tumorigenesis signals such as abnormal EGF signalling regulates MV release, remain unclear. Here, we set out to establish reliable readouts for MV biogenesis and then explore the molecular mechanisms that regulate MV generation. We found that Rho family small G protein Cdc42 is a convergent node of multiple regulatory signals that occur in MV biogenesis. The binding of activated GTP-bound Cdc42 and its downstream effector, Ras GTPase-activating-like protein 1 (IQGAP1), is required for MV shedding. Activated Cdc42 maintains sustained EGF signalling by inhibiting the internalization of cell surface receptors, including EGFR and the VEGF oligomer, VEGF90K, and then facilitates MV release. Subsequently, we further demonstrated that blocking these signalling pathways using the corresponding mutants effectively reduced MV shedding and significantly inhibited MV-promoted in vivo tumour angiogenesis. These findings reveal a complex regulation of MV shedding by tumour cells, shedding light on the regulatory mechanism of MV biogenesis, and potentially contributing to strategies that target MVs in cancer therapy.


Asunto(s)
Vesículas Extracelulares/metabolismo , Neoplasias/metabolismo , Microambiente Tumoral , Proteína de Unión al GTP cdc42/metabolismo , Células 3T3 , Animales , Comunicación Celular , Línea Celular , Línea Celular Tumoral , Micropartículas Derivadas de Células/metabolismo , Humanos , Ratones , Neovascularización Patológica/metabolismo , Transducción de Señal , Proteínas Activadoras de ras GTPasa/metabolismo
13.
J Biol Chem ; 284(27): 17914-25, 2009 Jul 03.
Artículo en Inglés | MEDLINE | ID: mdl-19403524

RESUMEN

Epidermal growth factor (EGF) exerts pleiotropic effects during oncogenesis, including the stimulation of cell migration and invasiveness. Although a number of traditional signaling proteins (e.g. Ras and Rho GTPases) have been implicated in EGF-stimulated cancer cell migration, less is known about the identity of those proteins functioning further downstream in this growth factor pathway. Here we have used HeLa carcinoma cells as a model system for investigating the role of tissue transglutaminase (TGase), a protein that has been linked to oncogenesis, in EGF-stimulated cancer cell migration and invasion. Treatment of HeLa cells with EGF resulted in TGase activation and its accumulation at their leading edges, whereas knocking down TGase expression, or treating cells with a TGase inhibitor, blocked EGF-stimulated cell migration and invasion. We show that EGF signaling through Ras and c-Jun N-terminal kinase is responsible for targeting TGase to the leading edges of cells and activating it. The requirement for EGF to properly localize and activate TGase can be circumvented by the expression of oncogenic Ras (G12V), whose ability to stimulate migration is also dependent on TGase. We further show that, in the highly aggressive breast cancer cell line MDAMB231, where EGF stimulation is unnecessary for migration and invasive activity, TGase is already at the leading edge and activated. These findings demonstrate that TGase plays a key role in cancer cell motility and invasiveness and represents a previously unappreciated participant in the EGF pathway that stimulates these processes in cancer cells.


Asunto(s)
Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Movimiento Celular/fisiología , Factor de Crecimiento Epidérmico/metabolismo , Transducción de Señal/fisiología , Transglutaminasas/metabolismo , Membrana Celular/enzimología , Supervivencia Celular/fisiología , Activación Enzimática/fisiología , Femenino , Proteínas de Unión al GTP , Genes ras/fisiología , Guanosina Trifosfato/metabolismo , Células HeLa , Humanos , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Invasividad Neoplásica , Proteína Glutamina Gamma Glutamiltransferasa 2 , ARN Interferente Pequeño , Transglutaminasas/genética
14.
PLoS One ; 15(4): e0231466, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-32298294

RESUMEN

DaHuangWan (DHW) is a traditional herbal medicine used by Mongolian to treat liver cancer for many years. Clinical application of the drug has been shown to help control tumor progression, prolong survival and improve quality of life. However, the underlying mechanisms and side effects of this drug remain unclear, which greatly limits the clinical application and further optimization of DHW. In this study, we found that DHW inhibits the proliferation of hepatoma cells by modulating the epithelial growth factor (EGF) signaling pathway. Berberine and Costunolide are the main active ingredients in DHW. Interestingly, the combination of Berberine and Costunolide has a dramatic synergistic effect on inhibiting the proliferation of hepatoma cells. Neither Berberine nor Costunolide directly block EGFR phosphorylation. Berberine promotes endocytosis of activated EGFR, while as Costunolide increases ubiquitination of EGFR and reduces EGFR recycling to cell membrane distribution, thereby inhibiting EGF signaling. Berberine and Costunolide target two different steps in regulating the EGF signaling, which explains the synergistic anti-cancer effect of DHW. Since Berberine and Costunolide do not directly target EGFR phosphorylation, DHW could be a supplementary medicine to tyrosine kinase inhibitors in cancer therapy.


Asunto(s)
Carcinoma Hepatocelular/tratamiento farmacológico , Factor de Crecimiento Epidérmico/antagonistas & inhibidores , Medicina de Hierbas/métodos , Neoplasias Hepáticas/tratamiento farmacológico , Medicina Tradicional de Asia Oriental/métodos , Apoptosis/efectos de los fármacos , Berberina/uso terapéutico , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Citometría de Flujo , Humanos , Mongolia , Plantas Medicinales , Sesquiterpenos/uso terapéutico
15.
Curr Biol ; 15(1): 1-10, 2005 Jan 11.
Artículo en Inglés | MEDLINE | ID: mdl-15649357

RESUMEN

BACKGROUND: Cloned-out of library-2 (Cool-2)/PAK-interactive exchange factor (alpha-Pix) was identified through its ability to bind the Cdc42/Rac target p21-activated kinase (PAK) and has been implicated in certain forms of X-linked mental retardation as well as in growth factor- and chemoattractant-coupled signaling pathways. We recently found that the dimeric form of Cool-2 is a specific guanine nucleotide exchange factor (GEF) for Rac, whereas monomeric Cool-2 is a GEF for Cdc42 as well as Rac. However, unlike many GEFs, Cool-2 binds to activated forms of Cdc42 and Rac. Thus, we have investigated the functional consequences of these interactions. RESULTS: We show that the binding of activated Cdc42 to the Cool-2 dimer markedly enhances its ability to associate with GDP bound Rac1, resulting in a significant activation of Rac-GEF activity. While the Rac-specific GEF activity of Cool-2 is mediated through the Dbl homology (DH) domain from one monomer and the Pleckstrin homology domain from the other, activated Cdc42 interacts with the DH domain, most likely opposite the DH domain binding site for GDP bound Rac. Activated Rac also binds to Cool-2; however, it strongly inhibits the GEF activity of dimeric Cool-2. CONCLUSIONS: We provide evidence for novel mechanisms of allosteric regulation of the Rac-GEF activity of the Cool-2 dimer, involving stimulatory effects by Cdc42 and feedback inhibition by Rac. These findings demonstrate that by serving as a target for GTP bound Cdc42 and a GEF for Rac, Cool-2 mediates a GTPase cascade where the activation of Cdc42 is translated into the activation of Rac.


Asunto(s)
Proteínas de Ciclo Celular/metabolismo , Factores de Intercambio de Guanina Nucleótido/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Transducción de Señal , Proteína de Unión al GTP cdc42/metabolismo , Proteínas de Unión al GTP rac/metabolismo , Animales , Células COS , Proteínas de Ciclo Celular/genética , Chlorocebus aethiops , Factores de Intercambio de Guanina Nucleótido/genética , Modelos Biológicos , Plásmidos/genética , Estructura Terciaria de Proteína , Factores de Intercambio de Guanina Nucleótido Rho , Quinasas p21 Activadas
16.
Artículo en Inglés | MEDLINE | ID: mdl-28670141

RESUMEN

Hyperglycemia or diabetes mellitus (DM), which is characterized by high blood glucose levels, has been linked to an increased risk of cancer for years. However, the underlying molecular mechanisms of the pathophysiological link are not yet fully understood. In this study, we demonstrate that high glucose levels promote the proliferation of breast cancer cells by stimulating epidermal growth factor receptor (EGFR) activation and the Rho family GTPase Rac1 and Cdc42 mediate the corresponding signaling induced by high glucose levels. We further show that Cdc42 promotes EGFR phosphorylation by blocking EGFR degradation, which may be mediated by the Cbl proteins, whereas the Rac1-mediated EGFR phosphorylation is independent of EGFR degradation. Our findings elucidate a part of the underlying molecular mechanism of the link between high glucose levels and tumorigenesis in breast cancer and may provide new insights on the therapeutic strategy for cancer patients with diabetes or hyperglycemia.

17.
Biol. Res ; 55: 35-35, 2022. ilus
Artículo en Inglés | LILACS | ID: biblio-1429901

RESUMEN

Extracellular vesicles (EVs) are naturally released membrane vesicles that act as carriers of proteins and RNAs for intercellular communication. With various biomolecules and specific ligands, EV has represented a novel form of information transfer, which possesses extremely outstanding efficiency and specificity compared to the classical signal transduction. In addition, EV has extended the concept of signal transduction to intercellular aspect by working as the collection of extracellular information. Therefore, the functions of EVs have been extensively characterized and EVs exhibit an exciting prospect for clinical applications. However, the biogenesis of EVs and, in particular, the regulation of this process by extracellular signals, which are essential to conduct further studies and support optimal utility, remain unclear. Here, we review the current understanding of the biogenesis of EVs, focus on the regulation of this process by extracellular signals and discuss their therapeutic value.


Asunto(s)
Vesículas Extracelulares/metabolismo , Transporte Biológico , ARN/metabolismo , Transducción de Señal , Comunicación Celular/fisiología
18.
Nat Commun ; 8: 14450, 2017 02 16.
Artículo en Inglés | MEDLINE | ID: mdl-28205552

RESUMEN

Non-classical secretory vesicles, collectively referred to as extracellular vesicles (EVs), have been implicated in different aspects of cancer cell survival and metastasis. Here, we describe how a specific class of EVs, called microvesicles (MVs), activates VEGF receptors and tumour angiogenesis through a unique 90 kDa form of VEGF (VEGF90K). We show that VEGF90K is generated by the crosslinking of VEGF165, catalysed by the enzyme tissue transglutaminase, and associates with MVs through its interaction with the chaperone Hsp90. We further demonstrate that MV-associated VEGF90K has a weakened affinity for Bevacizumab, causing Bevacizumab to be ineffective in blocking MV-dependent VEGF receptor activation. However, treatment with an Hsp90 inhibitor releases VEGF90K from MVs, restoring the sensitivity of VEGF90K to Bevacizumab. These findings reveal a novel mechanism by which cancer cell-derived MVs influence the tumour microenvironment and highlight the importance of recognizing their unique properties when considering drug treatment strategies.


Asunto(s)
Benzoquinonas/farmacología , Bevacizumab/farmacología , Neoplasias de la Mama/patología , Vesículas Extracelulares/clasificación , Vesículas Extracelulares/metabolismo , Lactamas Macrocíclicas/farmacología , Neovascularización Patológica/metabolismo , Receptores de Factores de Crecimiento Endotelial Vascular/metabolismo , Inductores de la Angiogénesis/metabolismo , Animales , Benzoquinonas/metabolismo , Bevacizumab/metabolismo , Línea Celular Tumoral/efectos de los fármacos , Línea Celular Tumoral/metabolismo , Micropartículas Derivadas de Células/clasificación , Micropartículas Derivadas de Células/metabolismo , Modelos Animales de Enfermedad , Combinación de Medicamentos , Femenino , Proteínas HSP90 de Choque Térmico/efectos de los fármacos , Proteínas HSP90 de Choque Térmico/metabolismo , Células HeLa , Humanos , Lactamas Macrocíclicas/metabolismo , Ratones , Neovascularización Patológica/patología , Vesículas Secretoras , Transducción de Señal , Transglutaminasas , Trasplante Heterólogo , Microambiente Tumoral , Factor A de Crecimiento Endotelial Vascular/efectos de los fármacos , Factor A de Crecimiento Endotelial Vascular/metabolismo
19.
Methods Enzymol ; 406: 58-69, 2006.
Artículo en Inglés | MEDLINE | ID: mdl-16472649

RESUMEN

The Cool (Cloned out of Library)/Pix (Pak interactive exchange factor) proteins have been implicated in a diversity of biological activities, ranging from pathways initiated by growth factors and chemoattractants to X-linked mental retardation. Initially discovered through yeast two-hybrid and biochemical analyses as binding partners for the Cdc42/Rac-target/effector, Pak (p21 activated kinase), the sequences for the Cool/Pix proteins revealed a DH (Dbl homology) domain. Because the DH domain is the limit functional unit for stimulating guanine nucleotide exchange on Rho family GTP-binding proteins, it was assumed that the Cool/Pix proteins would act as guanine nucleotide exchange factors (GEFs) for the Rho proteins. Of the three known isoforms, (p50Cool-1, p85Cool-1/beta-Pix, and 90Cool-2/alpha-Pix), only Cool-2/alpha-Pix has exhibited significant GEF activity. A number of experimental techniques have been used to characterize Cool-2, and in vitro analysis has revealed that its GEF activity is under tight control through intramolecular interactions involving several binding partners. Here we describe the biochemical methods used to study the Cool/Pix proteins and, in particular, the regulation of the GEF activity of Cool-2/alpha-Pix.


Asunto(s)
Proteínas de Ciclo Celular/metabolismo , Factores de Intercambio de Guanina Nucleótido/metabolismo , Animales , Células COS , Chlorocebus aethiops , Escherichia coli/metabolismo , Factores de Intercambio de Guanina Nucleótido/biosíntesis , Factores de Intercambio de Guanina Nucleótido/aislamiento & purificación , Guanosina Difosfato/metabolismo , Ratones , Células 3T3 NIH , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Recombinantes/aislamiento & purificación , Factores de Intercambio de Guanina Nucleótido Rho , Spodoptera , Proteína de Unión al GTP cdc42/biosíntesis , Proteína de Unión al GTP cdc42/aislamiento & purificación , Quinasas p21 Activadas , Proteínas de Unión al GTP rac/biosíntesis , Proteínas de Unión al GTP rac/aislamiento & purificación
20.
FEBS Lett ; 550(1-3): 119-23, 2003 Aug 28.
Artículo en Inglés | MEDLINE | ID: mdl-12935897

RESUMEN

Members of the Cool protein family contain SH3, Dbl, and pleckstrin homology domains and are binding partners for the p21-activated kinase (PAK). Using the yeast two-hybrid screen, we identified Cbl-b as a Cool family binding partner. We co-immunoprecipitated endogenous Cool and Cbl-b from a variety of breast cancer cell lines. The Cool-Cbl-b interaction requires the SH3 domain of Cool and competes with the binding of PAK to Cool proteins. Expression of Cbl-b effectively blocks the ability of Cool-2 to stimulate PAK, thus providing an additional mechanism, aside from catalyzing receptor ubiquitination, by which Cbl-b acts as a negative regulator for signaling activities requiring PAK activation.


Asunto(s)
Proteínas Adaptadoras Transductoras de Señales , Proteínas Portadoras/metabolismo , Proteínas de Ciclo Celular/metabolismo , Factores de Intercambio de Guanina Nucleótido/metabolismo , Fosfoproteínas/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Ubiquitina-Proteína Ligasas , Secuencia de Aminoácidos , Animales , Unión Competitiva , Neoplasias de la Mama/metabolismo , Células COS , Proteínas Portadoras/genética , Proteínas de Ciclo Celular/genética , Femenino , Factores de Intercambio de Guanina Nucleótido/genética , Humanos , Datos de Secuencia Molecular , Fosfoproteínas/genética , Unión Proteica , Proteínas Serina-Treonina Quinasas/genética , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas c-cbl , Factores de Intercambio de Guanina Nucleótido Rho , Células Tumorales Cultivadas , Técnicas del Sistema de Dos Híbridos , Quinasas p21 Activadas , Dominios Homologos src/fisiología
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