Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 14.623
Filtrar
1.
Int J Mol Sci ; 22(6)2021 Mar 18.
Artículo en Inglés | MEDLINE | ID: mdl-33803640

RESUMEN

The LATS1 kinase has been described as a tumor suppressor in various cancers. However, its role in melanoma has not been fully elucidated. There are several processes involved in tumorigenesis, including melanin production. Melanin content positively correlates with the level of reactive oxygen species (ROS) inside the cell. Accordingly, the purpose of the study was to assess the role of LATS1 in melanogenesis and oxidative stress and its influence on tumor growth. We have knocked down LATS1 in primary melanocytes and melanoma cells and found that its expression is crucial for melanin synthesis, ROS production, and oxidative stress response. We showed that LATS1 ablation significantly decreased the melanogenesis markers' expression and melanin synthesis in melanocyte and melanoma cell lines. Moreover, silencing LATS1 resulted in enhanced oxidative stress. Reduced melanin content in LATS1 knocked down tumors was associated with increased tumor growth, pointing to melanin's protective role in this process. The study demonstrated that LATS1 is highly engaged in melanogenesis and oxidative stress control and affects melanoma growth. Our results may find the implications in the diagnosis and treatment of pigmentation disorders, including melanoma.


Asunto(s)
Melaninas/biosíntesis , Melanoma/patología , Estrés Oxidativo , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Adulto , Animales , Biomarcadores de Tumor/metabolismo , Línea Celular Tumoral , Proliferación Celular , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Cinética , Melanocitos/metabolismo , Melanoma/genética , Ratones Desnudos , Especies Reactivas de Oxígeno/metabolismo , Estrés Fisiológico , Transactivadores/metabolismo , Factores de Transcripción/metabolismo , Hipoxia Tumoral/genética
2.
Nat Commun ; 12(1): 2424, 2021 04 23.
Artículo en Inglés | MEDLINE | ID: mdl-33893293

RESUMEN

Endocytosis mediates the cellular uptake of micronutrients and cell surface proteins. Fast Endophilin-mediated endocytosis, FEME, is not constitutively active but triggered upon receptor activation. High levels of growth factors induce spontaneous FEME, which can be suppressed upon serum starvation. This suggested a role for protein kinases in this growth factor receptor-mediated regulation. Using chemical and genetic inhibition, we find that Cdk5 and GSK3ß are negative regulators of FEME. They antagonize the binding of Endophilin to Dynamin-1 and to CRMP4, a Plexin A1 adaptor. This control is required for proper axon elongation, branching and growth cone formation in hippocampal neurons. The kinases also block the recruitment of Dynein onto FEME carriers by Bin1. As GSK3ß binds to Endophilin, it imposes a local regulation of FEME. Thus, Cdk5 and GSK3ß are key regulators of FEME, licensing cells for rapid uptake by the pathway only when their activity is low.


Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/genética , Quinasa 5 Dependiente de la Ciclina/genética , Endocitosis/genética , Glucógeno Sintasa Quinasa 3 beta/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Animales , Células Cultivadas , Clatrina/metabolismo , Quinasa 5 Dependiente de la Ciclina/metabolismo , Dinamina I/genética , Dinamina I/metabolismo , Regulación de la Expresión Génica , Glucógeno Sintasa Quinasa 3 beta/metabolismo , Células HEK293 , Células HeLa , Hipocampo/citología , Hipocampo/metabolismo , Humanos , Ratones Endogámicos C57BL , Proteínas Musculares/genética , Proteínas Musculares/metabolismo , Neuronas/metabolismo , Unión Proteica , Interferencia de ARN
3.
Medicine (Baltimore) ; 100(14): e25278, 2021 Apr 09.
Artículo en Inglés | MEDLINE | ID: mdl-33832094

RESUMEN

ABSTRACT: A kinase interacting protein 1 (AKIP1) is upregulated in cancer cells/tissues and associated with deteriorative tumor features, while it has not been investigated in tongue squamous cell carcinoma (TSCC). The goal of this study was to measure AKIP1 expression and analyze its correlation with clinical feature and prognosis in TSCC patients.We retrospectively reviewed 194 TSCC patients, whose formalin fixed paraffin-embedded (FFPE) tumor tissue specimens and paired adjacent tissue specimens were accessible for AKIP1 detection by immunohistochemistry (IHC). Whereas only 107 patients whose fresh-frozen tumor tissue and paired fresh-frozen adjacent tissue that were still available in storage were included for AKIP1 mRNA detection by real-time reverse transcription quantitative polymerase chain reaction (RT-qPCR).AKIP1 expression (both the protein detected by IHC and mRNA detected by RT-qPCR) was higher in TSCC tissue than that in adjacent tissue. In addition, both tumor AKIP1 mRNA and protein expressions were correlated with advanced N stage and TNM stage, while they were not correlated with other clinical features in TSCC patients. As for survival, there was a correlation of AKIP1 mRNA with poor overall survival (OS), while the correlation of AKIP1 protein expression with OS was of limited statistical significance.There is an upregulation of AKIP1 in TSCC and it correlates with lymph node metastasis as well as unfavorable prognosis in TSCC patients.


Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Carcinoma de Células Escamosas/genética , Proteínas Nucleares/metabolismo , Neoplasias de la Lengua/genética , Regulación hacia Arriba , Adulto , Anciano , Biomarcadores/análisis , Carcinoma de Células Escamosas/patología , Estudios de Casos y Controles , Supervivencia sin Enfermedad , Humanos , Metástasis Linfática/genética , Persona de Mediana Edad , ARN Mensajero/metabolismo , Estudios Retrospectivos , Neoplasias de la Lengua/patología
4.
Nat Commun ; 12(1): 2299, 2021 04 16.
Artículo en Inglés | MEDLINE | ID: mdl-33863908

RESUMEN

Mycobacterial cell-wall glycolipids elicit an anti-mycobacterial immune response via FcRγ-associated C-type lectin receptors, including Mincle, and caspase-recruitment domain family member 9 (CARD9). Additionally, mycobacteria harbor immuno-evasive cell-wall lipids associated with virulence and latency; however, a mechanism of action is unclear. Here, we show that the DAP12-associated triggering receptor expressed on myeloid cells 2 (TREM2) recognizes mycobacterial cell-wall mycolic acid (MA)-containing lipids and suggest a mechanism by which mycobacteria control host immunity via TREM2. Macrophages respond to glycosylated MA-containing lipids in a Mincle/FcRγ/CARD9-dependent manner to produce inflammatory cytokines and recruit inducible nitric oxide synthase (iNOS)-positive mycobactericidal macrophages. Conversely, macrophages respond to non-glycosylated MAs in a TREM2/DAP12-dependent but CARD9-independent manner to recruit iNOS-negative mycobacterium-permissive macrophages. Furthermore, TREM2 deletion enhances Mincle-induced macrophage activation in vitro and inflammation in vivo and accelerates the elimination of mycobacterial infection, suggesting that TREM2-DAP12 signaling counteracts Mincle-FcRγ-CARD9-mediated anti-mycobacterial immunity. Mycobacteria, therefore, harness TREM2 for immune evasion.


Asunto(s)
Evasión Inmune , Tuberculosis Latente/inmunología , Glicoproteínas de Membrana/metabolismo , Mycobacterium tuberculosis/inmunología , Ácidos Micólicos/metabolismo , Receptores Inmunológicos/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Animales , Proteínas Adaptadoras de Señalización CARD/genética , Proteínas Adaptadoras de Señalización CARD/metabolismo , Pared Celular/metabolismo , Células Cultivadas , Modelos Animales de Enfermedad , Femenino , Glucolípidos/metabolismo , Humanos , Tuberculosis Latente/microbiología , Lectinas Tipo C/genética , Lectinas Tipo C/metabolismo , Activación de Macrófagos/inmunología , Macrófagos/inmunología , Masculino , Glicoproteínas de Membrana/genética , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Ratones , Ratones Noqueados , Mycobacterium tuberculosis/metabolismo , Mycobacterium tuberculosis/patogenicidad , Cultivo Primario de Células , Receptores de IgG/metabolismo , Receptores Inmunológicos/genética , Factores de Virulencia/metabolismo
5.
Int J Mol Sci ; 22(5)2021 Feb 25.
Artículo en Inglés | MEDLINE | ID: mdl-33669042

RESUMEN

Since its first identification as a cardiac transverse tubule (t-tubule) protein, followed by the cloning of the cardiac isoform responsible for t-tubule membrane microdomain formation, cardiac bridging integrator 1 (cBIN1) and its organized microdomains have emerged as a key mechanism in maintaining normal beat-to-beat heart contraction and relaxation. The abnormal remodeling of cBIN1-microdomains occurs in stressed and diseased cardiomyocytes, contributing to the pathophysiology of heart failure. Due to the homeostatic turnover of t-tubule cBIN1-microdomains via microvesicle release into the peripheral circulation, plasma cBIN1 can be assayed as a liquid biopsy of cardiomyocyte health. A new blood test cBIN1 score (CS) has been developed as a dimensionless inverse index derived from plasma cBIN1 concentration with a diagnostic and prognostic power for clinical outcomes in stable ambulatory patients with heart failure with reduced or preserved ejection fraction (HFrEF or HFpEF). Recent evidence further indicates that exogenous cBIN1 introduced by adeno-associated virus 9-based gene therapy can rescue cardiac contraction and relaxation in failing hearts. The therapeutic potential of cBIN1 gene therapy is enormous given its ability to rescue cardiac inotropy and provide lusitropic protection in the meantime. These unprecedented capabilities of cBIN1 gene therapy are shifting the current paradigm of therapy development for heart failure, particularly HFpEF.


Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/sangre , Terapia Genética/métodos , Insuficiencia Cardíaca/sangre , Miocitos Cardíacos/metabolismo , Proteínas Nucleares/sangre , Retículo Sarcoplasmático/metabolismo , Proteínas Supresoras de Tumor/sangre , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Animales , Biomarcadores/sangre , Señalización del Calcio/fisiología , Insuficiencia Cardíaca/diagnóstico , Insuficiencia Cardíaca/metabolismo , Insuficiencia Cardíaca/fisiopatología , Humanos , Proteínas de la Membrana/metabolismo , Contracción Miocárdica , Miocitos Cardíacos/patología , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Dominios Proteicos , Sarcolema/metabolismo , Retículo Sarcoplasmático/patología , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismo
7.
Nat Cell Biol ; 23(3): 257-267, 2021 03.
Artículo en Inglés | MEDLINE | ID: mdl-33723425

RESUMEN

The complexity of intracellular signalling requires both a diversity of molecular players and the sequestration of activity to unique compartments within the cell. Recent findings on the role of liquid-liquid phase separation provide a distinct mechanism for the spatial segregation of proteins to regulate signalling pathway crosstalk. Here, we discover that DACT1 is induced by TGFß and forms protein condensates in the cytoplasm to repress Wnt signalling. These condensates do not localize to any known organelles but, rather, exist as phase-separated proteinaceous cytoplasmic bodies. The deletion of intrinsically disordered domains within the DACT1 protein eliminates its ability to both form protein condensates and suppress Wnt signalling. Isolation and mass spectrometry analysis of these particles revealed a complex of protein machinery that sequesters casein kinase 2-a Wnt pathway activator. We further demonstrate that DACT1 condensates are maintained in vivo and that DACT1 is critical to breast and prostate cancer bone metastasis.


Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Neoplasias Óseas/metabolismo , Neoplasias de la Mama/metabolismo , Movimiento Celular/efectos de los fármacos , Proteínas Nucleares/metabolismo , Neoplasias de la Próstata/metabolismo , Factor de Crecimiento Transformador beta/farmacología , Vía de Señalización Wnt/efectos de los fármacos , Proteína Wnt3A/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Animales , Neoplasias Óseas/genética , Neoplasias Óseas/secundario , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Quinasa de la Caseína II/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Femenino , Regulación Neoplásica de la Expresión Génica , Células HEK293 , Humanos , Masculino , Ratones Endogámicos NOD , Ratones Desnudos , Ratones SCID , Invasividad Neoplásica , Proteínas Nucleares/genética , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/patología , Proteína Wnt3A/genética
8.
Int J Mol Sci ; 22(5)2021 Feb 25.
Artículo en Inglés | MEDLINE | ID: mdl-33668983

RESUMEN

Transcytosis of polymeric IgA and IgM from the basolateral surface to the apical side of the epithelium and subsequent secretion into mucosal fluids are mediated by the polymeric immunoglobulin receptor (pIgR). Secreted IgA and IgM have vital roles in mucosal immunity in response to pathogenic infections. Binding and recognition of polymeric IgA and IgM by pIgR require the joining chain (J chain), a small protein essential in the formation and stabilization of polymeric Ig structures. Recent studies have identified marginal zone B and B1 cell-specific protein (MZB1) as a novel regulator of polymeric IgA and IgM formation. MZB1 might facilitate IgA and IgM transcytosis by promoting the binding of J chain to Ig. In this review, we discuss the roles of pIgR in transcytosis of IgA and IgM, the roles of J chain in the formation of polymeric IgA and IgM and recognition by pIgR, and focus particularly on recent progress in understanding the roles of MZB1, a molecular chaperone protein.


Asunto(s)
Inmunoglobulina A/metabolismo , Inmunoglobulina M/metabolismo , Receptores de Inmunoglobulina Polimérica/metabolismo , Transcitosis , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Animales , Humanos , Polimerizacion
9.
Nat Commun ; 12(1): 1716, 2021 03 19.
Artículo en Inglés | MEDLINE | ID: mdl-33741917

RESUMEN

Cancer stem cells (CSCs) are a small but critical cell population for cancer biology since they display inherent resistance to standard therapies and give rise to metastases. Despite accruing evidence establishing a link between deregulation of epitranscriptome-related players and tumorigenic process, the role of messenger RNA (mRNA) modifications in the regulation of CSC properties remains poorly understood. Here, we show that the cytoplasmic pool of fat mass and obesity-associated protein (FTO) impedes CSC abilities in colorectal cancer through its N6,2'-O-dimethyladenosine (m6Am) demethylase activity. While m6Am is strategically located next to the m7G-mRNA cap, its biological function is not well understood and has not been addressed in cancer. Low FTO expression in patient-derived cell lines elevates m6Am level in mRNA which results in enhanced in vivo tumorigenicity and chemoresistance. Inhibition of the nuclear m6Am methyltransferase, PCIF1/CAPAM, fully reverses this phenotype, stressing the role of m6Am modification in stem-like properties acquisition. FTO-mediated regulation of m6Am marking constitutes a reversible pathway controlling CSC abilities. Altogether, our findings bring to light the first biological function of the m6Am modification and its potential adverse consequences for colorectal cancer management.


Asunto(s)
Dioxigenasa FTO Dependiente de Alfa-Cetoglutarato/metabolismo , Neoplasias Colorrectales/metabolismo , Citoplasma/metabolismo , Desmetilación , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Adenosina/metabolismo , Dioxigenasa FTO Dependiente de Alfa-Cetoglutarato/genética , Línea Celular Tumoral , Núcleo Celular/metabolismo , Neoplasias Colorrectales/genética , Regulación Neoplásica de la Expresión Génica , Silenciador del Gen , Humanos , Metiltransferasas/metabolismo , Proteínas Nucleares/metabolismo , ARN Mensajero/metabolismo
10.
Nat Commun ; 12(1): 1758, 2021 03 19.
Artículo en Inglés | MEDLINE | ID: mdl-33741948

RESUMEN

The molecular machinery and chromosome structures carrying out meiosis are frequently conserved from yeast to mammals. However, signals initiating meiosis appear divergent: while nutrient restriction induces meiosis in the yeast system, retinoic acid (RA) and its target Stra8 have been shown to be necessary but not sufficient to induce meiotic initiation in mammalian germ cells. Here, we use primary culture of mouse undifferentiated spermatogonia without the support of gonadal somatic cells to show that nutrient restriction in combination with RA is sufficient to induce Stra8- and Spo11-dependent meiotic gene and chromosome programs that recapitulate the transcriptomic and cytologic features of in vivo meiosis. We demonstrate that neither nutrient restriction nor RA alone exerts these effects. Moreover, we identify a distinctive network of 11 nutrient restriction-upregulated transcription factor genes, which are associated with early meiosis in vivo and whose expression does not require RA. Our study proposes a conserved model, in which nutrient restriction induces meiotic initiation by upregulating key transcription factor genes for the meiotic gene program and provides an in vitro platform for meiotic induction that could facilitate research and haploid gamete production.


Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Endodesoxirribonucleasas/metabolismo , Meiosis/efectos de los fármacos , Nutrientes/metabolismo , Espermatogonias/efectos de los fármacos , Tretinoina/farmacología , Proteínas Adaptadoras Transductoras de Señales/genética , Animales , Células Cultivadas , Endodesoxirribonucleasas/genética , Expresión Génica/efectos de los fármacos , Masculino , Mamíferos/genética , Mamíferos/metabolismo , Meiosis/genética , Ratones Endogámicos C57BL , Ratones Endogámicos DBA , Ratones Noqueados , Análisis de Secuencia de ARN/métodos , Análisis de la Célula Individual/métodos , Espermatogonias/citología , Espermatogonias/metabolismo
11.
Int J Mol Sci ; 22(4)2021 Feb 18.
Artículo en Inglés | MEDLINE | ID: mdl-33670622

RESUMEN

The Hippo pathway is involved in human tumorigenesis and tissue repair. Here, we investigated the Hippo coactivator Yes-associated protein 1 (YAP1) and the kinase large tumor suppressor 1/2 (LATS1/2) in tumors of the parathyroid glands, which are almost invariably associated with primary hyperparathyroidism. Compared with normal parathyroid glands, parathyroid adenomas (PAds) and carcinomas show variably but reduced nuclear YAP1 expression. The kinase LATS1/2, which phosphorylates YAP1 thus promoting its degradation, was also variably reduced in PAds. Further, YAP1 silencing reduces the expression of the key parathyroid oncosuppressor multiple endocrine neoplasia type 1(MEN1), while MEN1 silencing increases YAP1 expression. Treatment of patient-derived PAds-primary cell cultures and Human embryonic kidney 293A (HEK293A) cells expressing the calcium-sensing receptor (CASR) with the CASR agonist R568 induces YAP1 nuclear accumulation. This effect was prevented by the incubation of the cells with RhoA/Rho-associated coiled-coil-containing protein kinase (ROCK) inhibitors Y27632 and H1152. Lastly, CASR activation increased the expression of the YAP1 gene targets CYR61, CTGF, and WNT5A, and this effect was blunted by YAP1 silencing. Concluding, here we provide preliminary evidence of the involvement of the Hippo pathway in human tumor parathyroid cells and of the existence of a CASR-ROCK-YAP1 axis. We propose a tumor suppressor role for YAP1 and LATS1/2 in parathyroid tumors.


Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/genética , Glándulas Paratiroides/metabolismo , Neoplasias de las Paratiroides/genética , Receptores Sensibles al Calcio/genética , Factores de Transcripción/genética , 1-(5-Isoquinolinesulfonil)-2-Metilpiperazina/análogos & derivados , 1-(5-Isoquinolinesulfonil)-2-Metilpiperazina/farmacología , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Amidas/farmacología , Núcleo Celular/efectos de los fármacos , Núcleo Celular/metabolismo , Expresión Génica/efectos de los fármacos , Células HEK293 , Humanos , Neoplasias de las Paratiroides/metabolismo , Fenetilaminas/farmacología , Propilaminas/farmacología , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas/metabolismo , Piridinas/farmacología , Interferencia de ARN , Receptores Sensibles al Calcio/agonistas , Receptores Sensibles al Calcio/metabolismo , Factores de Transcripción/metabolismo , Células Tumorales Cultivadas , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismo , Quinasas Asociadas a rho/antagonistas & inhibidores , Quinasas Asociadas a rho/metabolismo
12.
Int J Mol Sci ; 22(4)2021 Feb 14.
Artículo en Inglés | MEDLINE | ID: mdl-33672928

RESUMEN

Extensive water loss and melanin hyperproduction can cause various skin disorders. Low-temperature argon plasma (LTAP) has shown the possibility of being used for the treatment of various skin diseases, such as atopic dermatitis and skin cancer. However, the role of LTAP in regulating skin moisturizing and melanogenesis has not been investigated. In this study, we aimed to determine the effect of LTAP on yes-associated protein (YAP), a major transcriptional coactivator in the Hippo signaling pathway that is involved in skin moisturizing and melanogenesis-regulating markers. In normal human epidermal keratinocytes (NHEKs), the human epidermal keratinocyte line HaCaT, and human dermal fibroblasts (HDFs), we found that LTAP exhibited increased expression levels of YAP protein. In addition, the expression levels of filaggrin (FLG), which is involved in natural moisturizing factors (NMFs), and hyaluronic acid synthase (HAS), transglutaminase (TGM), and involucrin (IVL), which regulate skin barrier and moisturizing, were also increased after exposure to LTAP. Furthermore, collagen type I alpha 1 and type III alpha 1 (COL1A1, COL3A1) were increased after LTAP exposure, but the expression level of matrix metalloproteinase-3 (MMP-3) was reduced. Moreover, LTAP was found to suppress alpha-melanocyte stimulating hormone (α-MSH)-induced melanogenesis in murine melanoma B16F10 cells and normal human melanocytes (NHEMs). LTAP regulates melanogenesis of the melanocytes through decreased YAP pathway activation in a melanocortin 1 receptor (MC1R)-dependent manner. Taken together, our data show that LTAP regulates skin moisturizing and melanogenesis through modulation of the YAP pathway, and the effect of LTAP on the expression level of YAP varies from cell to cell. Thus, LTAP might be developed as a treatment method to improve the skin barrier, moisture content, and wrinkle formation, and to reduce melanin generation.


Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Argón/farmacología , Melaninas/metabolismo , Gases em Plasma/farmacología , Piel/efectos de los fármacos , Factores de Transcripción/metabolismo , Animales , Línea Celular , Línea Celular Tumoral , Células Cultivadas , Humanos , Queratinocitos/citología , Queratinocitos/efectos de los fármacos , Queratinocitos/metabolismo , Melanocitos/citología , Melanocitos/efectos de los fármacos , Melanocitos/metabolismo , Ratones , Receptor de Melanocortina Tipo 1/metabolismo , Piel/metabolismo , Envejecimiento de la Piel/efectos de los fármacos , Temperatura , alfa-MSH/metabolismo
13.
Int J Mol Sci ; 22(4)2021 Feb 12.
Artículo en Inglés | MEDLINE | ID: mdl-33673180

RESUMEN

Bam32 (B cell adaptor molecule of 32 kDa) functions in the immune responses of various leukocytes. However, the role of neutrophil Bam32 in inflammation is entirely unknown. Here, we determined the role of Bam32 in chemokine CXCL2-induced neutrophil chemotaxis in three mouse models of neutrophil recruitment. By using intravital microscopy in the mouse cremaster muscle, we found that transmigrated neutrophil number, neutrophil chemotaxis velocity, and total neutrophil chemotaxis distance were increased in Bam32-/- mice when compared with wild-type (WT) mice. In CXCL2-induced mouse peritonitis, the total emigrated neutrophils were increased in Bam32-/- mice at 2 but not 4 h. The CXCL2-induced chemotaxis distance and migration velocity of isolated Bam32-/- neutrophils in vitro were increased. We examined the activation of small GTPases Rac1, Rac2, and Rap1; the levels of phospho-Akt2 and total Akt2; and their crosstalk with Bam32 in neutrophils. The deficiency of Bam32 suppressed Rap1 activation without changing the activation of Rac1 and Rac2. The pharmacological inhibition of Rap1 by geranylgeranyltransferase I inhibitor (GGTI298) increased WT neutrophil chemotaxis. In addition, the deficiency of Bam32, as well as the inhibition of Rap1 activation, increased the levels of CXCL2-induced Akt1/2 phosphorylation at Thr308/309 in neutrophils. The inhibition of Akt by SH-5 attenuated CXCL2-induced adhesion and emigration in Bam32-/- mice. Together, our results reveal that Bam32 has a suppressive role in chemokine-induced neutrophil chemotaxis by regulating Rap1 activation and that this role of Bam32 in chemokine-induced neutrophil recruitment relies on the activation of PI3K effector Akt.


Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Quimiocinas/metabolismo , Lipoproteínas/metabolismo , Infiltración Neutrófila , Neutrófilos/metabolismo , Transducción de Señal , Proteínas Adaptadoras Transductoras de Señales/genética , Animales , Quimiocinas/genética , Lipoproteínas/genética , Ratones , Ratones Noqueados , Fosfatidilinositol 3-Quinasas/genética , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/genética , Proteínas Proto-Oncogénicas c-akt/metabolismo , Proteínas de Unión al GTP rho/genética , Proteínas de Unión al GTP rho/metabolismo
14.
Front Immunol ; 12: 573078, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33692778

RESUMEN

Swine acute diarrhea syndrome coronavirus (SADS-CoV), first discovered in 2017, is a porcine enteric coronavirus that can cause acute diarrhea syndrome (SADS) in piglets. Here, we studied the role of SADS-CoV nucleocapsid (N) protein in innate immunity. Our results showed that SADS-CoV N protein could inhibit type I interferon (IFN) production mediated by Sendai virus (Sev) and could block the phosphorylation and nuclear translocation of interferon regulatory factor 3 (IRF3). Simultaneously, the IFN-ß promoter activity mediated by TANK binding kinase 1 (TBK1) or its upstream molecules in the RLRs signal pathway was inhibited by SADS-CoV N protein. Further investigations revealed that SADS-CoV N protein could counteract interaction between TNF receptor-associated factor 3 (TRAF3) and TBK1, which led to reduced TBK1 activation and IFN-ß production. Our study is the first report of the interaction between SADS-CoV N protein and the host antiviral innate immune responses, and the mechanism utilized by SADS-CoV N protein provides a new insight of coronaviruses evading host antiviral innate immunity.


Asunto(s)
Alphacoronavirus/metabolismo , Interferón beta/antagonistas & inhibidores , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Factor 3 Asociado a Receptor de TNF/antagonistas & inhibidores , Proteínas Adaptadoras Transductoras de Señales/inmunología , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Alphacoronavirus/inmunología , Animales , Línea Celular , Coronavirus/inmunología , Coronavirus/metabolismo , Células HEK293 , Interacciones Huésped-Patógeno , Humanos , Quinasa I-kappa B/inmunología , Quinasa I-kappa B/metabolismo , Inmunidad Innata , Factor 3 Regulador del Interferón/inmunología , Factor 3 Regulador del Interferón/metabolismo , Interferón beta/biosíntesis , Interferón beta/inmunología , Interferón beta/metabolismo , FN-kappa B/inmunología , FN-kappa B/metabolismo , Proteínas Serina-Treonina Quinasas/inmunología , Proteínas Serina-Treonina Quinasas/metabolismo , Porcinos , Factor 3 Asociado a Receptor de TNF/inmunología , Factor 3 Asociado a Receptor de TNF/metabolismo
15.
Molecules ; 26(5)2021 Feb 25.
Artículo en Inglés | MEDLINE | ID: mdl-33668689

RESUMEN

Rapamycin is an antifungal drug with antitumor activity and acts inhibiting the mTOR complex. Due to drug antitumor potential, the aim of this study was to evaluate its effect on a preclinical model of primary mammary gland tumors and their metastases from female dogs. Four cell lines from our cell bank, two from primary canine mammary tumors (UNESP-CM1, UNESP-CM60) and two metastases (UNESP-MM1, and UNESP-MM4) were cultured in vitro and investigated for rapamycin IC50. Then, cell lines were treated with rapamycin IC50 dose and mRNA and protein were extracted in treated and non-treated cells to perform AKT, mTOR, PTEN and 4EBP1 gene expression and global proteomics by mass spectrometry. MTT assay demonstrated rapamycin IC50 dose for all different tumor cells between 2 and 10 µM. RT-qPCR from cultured cells, control versus treated group and primary tumor cells versus metastatic tumor cells, did not shown statistical differences. In proteomics were found 273 proteins in all groups, and after data normalization 49 and 92 proteins were used for statistical analysis for comparisons between control versus rapamycin treatment groups, and metastasis versus primary tumor versus metastasis rapamycin versus primary tumor rapamycin, respectively. Considering the two statistical analysis, four proteins, phosphoglycerate mutase, malate dehydrogenase, l-lactate dehydrogenase and nucleolin were found in decreased abundance in the rapamycin group and they are related with cellular metabolic processes and enhanced tumor malignant behavior. Two proteins, dihydrolipoamide dehydrogenase and superoxide dismutase, also related with metabolic processes, were found in higher abundance in rapamycin group and are associated with apoptosis. The results suggested that rapamycin was able to inhibit cell growth of mammary gland tumor and metastatic tumors cells in vitro, however, concentrations needed to reach the IC50 were higher when compared to other studies.


Asunto(s)
Antibióticos Antineoplásicos/farmacología , Neoplasias Mamarias Animales/tratamiento farmacológico , Proteómica , Sirolimus/farmacología , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Animales , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Perros , Ensayos de Selección de Medicamentos Antitumorales , Femenino , Neoplasias Mamarias Animales/metabolismo , Neoplasias Mamarias Animales/patología , Espectrometría de Masas , Fosfohidrolasa PTEN/genética , Fosfohidrolasa PTEN/metabolismo , Proteínas Proto-Oncogénicas c-akt/genética , Proteínas Proto-Oncogénicas c-akt/metabolismo , Serina-Treonina Quinasas TOR/genética , Serina-Treonina Quinasas TOR/metabolismo , Células Tumorales Cultivadas
16.
Cell Prolif ; 54(5): e13023, 2021 May.
Artículo en Inglés | MEDLINE | ID: mdl-33759281

RESUMEN

BACKGROUND: Vascular smooth muscle cells (VSMC) switch to macrophage-like cells after cholesterol loading, and this change may play an important role in atherogenesis. Muscleblind-like splicing regulator 1 (MBNL1) is a well-known splicing factor that has been implicated in many cellular processes. However, the role of MBNL1 in VSMC macrophage-like transdifferentiation is largely unknown. In this study, we aim to characterize the role of MBNL1-induced gene splicing during atherogenesis. METHODS: The expression of MBNL1 and Abelson interactor 1 (Abi1) splice variants (Abi1-e10 and Abi1-Δe10) was compared between artery tissues from healthy donors and atherosclerosis patients. Regulatory mechanisms of MBNL1-induced Abi1 gene splicing were studied, and the signal pathways mediated by Abi1 splice variants were investigated in VSMC. RESULTS: Loss of MBNL1 was found in the macrophage-like VSMC (VSMC-M) in artery wall from atherosclerosis patients. In vitro and in vivo evidence confirmed that Abi1 is one of the MBNL1 target genes. Loss of MBNL1 significantly induces the Abi1-Δe10 isoform expression. Compared to the known actin organization activities of the Abi1 gene, we discovered a novel action of Abi1-Δe10, whereby Abi1-Δe10 activates Rac1 independent of upstream stimulation and triggers the Rac1-NOX1-ROS pathway, which results in increased expression of transcription factor Kruppel-like factor 4 (KLF4). While Abi1-Δe10 inhibits contractile VSMC biomarkers expression and cell contraction, it stimulates VSMC proliferation, migration and macrophage-like transdifferentiation. CONCLUSION: Loss-of-function of MBNL1 activates VSMC-M transdifferentiation to promote atherogenesis through regulating Abi1 RNA splicing.


Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas del Citoesqueleto/genética , Músculo Liso Vascular/metabolismo , Empalme del ARN , Proteínas de Unión al ARN/metabolismo , Actinas/metabolismo , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Aterosclerosis/metabolismo , Aterosclerosis/patología , Desdiferenciación Celular , Células Cultivadas , Proteínas del Citoesqueleto/metabolismo , Humanos , Factores de Transcripción de Tipo Kruppel/metabolismo , Macrófagos/citología , Macrófagos/metabolismo , Músculo Liso Vascular/citología , NADPH Oxidasa 1/antagonistas & inhibidores , NADPH Oxidasa 1/genética , NADPH Oxidasa 1/metabolismo , Fenotipo , Isoformas de Proteínas/metabolismo , Interferencia de ARN , ARN Interferente Pequeño/metabolismo , Proteínas de Unión al ARN/antagonistas & inhibidores , Proteínas de Unión al ARN/genética , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal , Proteína de Unión al GTP rac1/metabolismo
17.
Am J Physiol Renal Physiol ; 320(5): F734-F747, 2021 05 01.
Artículo en Inglés | MEDLINE | ID: mdl-33682442

RESUMEN

The physiological role of the shorter isoform of with no lysine kinase (WNK)1 that is exclusively expressed in the kidney (KS-WNK1), with particular abundance in the distal convoluted tubule, remains elusive. KS-WNK1, despite lacking the kinase domain, is nevertheless capable of stimulating the NaCl cotransporter, apparently through activation of WNK4. It has recently been shown that a less severe form of familial hyperkalemic hypertension featuring only hyperkalemia is caused by missense mutations in the WNK1 acidic domain that preferentially affect cullin 3 (CUL3)-Kelch-like protein 3 (KLHL3) E3-induced degradation of KS-WNK1 rather than that of full-length WNK1. Here, we show that full-length WNK1 is indeed less impacted by the CUL3-KLHL3 E3 ligase complex compared with KS-WNK1. We demonstrated that the unique 30-amino acid NH2-terminal fragment of KS-WNK1 is essential for its activating effect on the NaCl cotransporter and recognition by KLHL3. We identified specific amino acid residues in this region critical for the functional effect of KS-WNK1 and KLHL3 sensitivity. To further explore this, we generated KLHL3-R528H knockin mice that mimic human mutations causing familial hyperkalemic hypertension. These mice revealed that the KLHL3 mutation specifically increased expression of KS-WNK1 in the kidney. We also observed that in wild-type mice, the expression of KS-WNK1 was only detectable after exposure to a low-K+ diet. These findings provide new insights into the regulation and function of KS-WNK1 by the CUL3-KLHL3 complex in the distal convoluted tubule and indicate that this pathway is regulated by dietary K+ levels.NEW & NOTEWORTHY In this work, we demonstrated that the kidney-specific isoform of with no lysine kinase 1 (KS-WNK1) in the kidney is modulated by dietary K+ and activity of the ubiquitin ligase protein Kelch-like protein 3. We analyzed the role of different amino acid residues of KS-WNK1 in its activity against the NaCl cotransporter and sensitivity to Kelch-like protein 3.


Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Riñón/enzimología , Proteínas de Microfilamentos/metabolismo , Potasio en la Dieta/metabolismo , Seudohipoaldosteronismo/enzimología , Proteína Quinasa Deficiente en Lisina WNK 1/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Animales , Proteínas Cullin/metabolismo , Estabilidad de Enzimas , Femenino , Riñón/fisiopatología , Ratones Endogámicos C57BL , Ratones Transgénicos , Proteínas de Microfilamentos/genética , Mutación , Dominios y Motivos de Interacción de Proteínas , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Proteolisis , Seudohipoaldosteronismo/genética , Seudohipoaldosteronismo/fisiopatología , Miembro 3 de la Familia de Transportadores de Soluto 12/genética , Miembro 3 de la Familia de Transportadores de Soluto 12/metabolismo , Proteína Quinasa Deficiente en Lisina WNK 1/deficiencia , Proteína Quinasa Deficiente en Lisina WNK 1/genética , Xenopus laevis
18.
Nat Commun ; 12(1): 1648, 2021 03 12.
Artículo en Inglés | MEDLINE | ID: mdl-33712605

RESUMEN

Cardiomyocytes undergo significant structural and functional changes after birth, and these fundamental processes are essential for the heart to pump blood to the growing body. However, due to the challenges of isolating single postnatal/adult myocytes, how individual newborn cardiomyocytes acquire multiple aspects of the mature phenotype remains poorly understood. Here we implement large-particle sorting and analyze single myocytes from neonatal to adult hearts. Early myocytes exhibit wide-ranging transcriptomic and size heterogeneity that is maintained until adulthood with a continuous transcriptomic shift. Gene regulatory network analysis followed by mosaic gene deletion reveals that peroxisome proliferator-activated receptor coactivator-1 signaling, which is active in vivo but inactive in pluripotent stem cell-derived cardiomyocytes, mediates the shift. This signaling simultaneously regulates key aspects of cardiomyocyte maturation through previously unrecognized proteins, including YAP1 and SF3B2. Our study provides a single-cell roadmap of heterogeneous transitions coupled to cellular features and identifies a multifaceted regulator controlling cardiomyocyte maturation.


Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Miocitos Cardíacos/metabolismo , Receptores Activados del Proliferador del Peroxisoma/metabolismo , Factores de Empalme de ARN/metabolismo , Factores de Transcripción/metabolismo , Animales , Calcio/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Diferenciación Celular , Redes Reguladoras de Genes , Humanos , Células Madre Pluripotentes Inducidas/metabolismo , Ratones , Receptores Activados del Proliferador del Peroxisoma/genética , Células Madre Pluripotentes/metabolismo , Transducción de Señal , Factores de Transcripción/genética , Transcriptoma
19.
Mol Med Rep ; 23(6)2021 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-33786612

RESUMEN

Dioscin, an extract from traditional Chinese herbal plants, displays various biological and pharmacological effects on tumors, including inhibition of cell proliferation and induction of DNA damage. However, the effects of dioscin on oral squamous cell carcinoma (OSCC) cells are not completely understood. The present study aimed to evaluate the impact of dioscin on OSCC cell proliferation. Cell Counting Kit­8 and 5­ethynyl­2'­deoxyuridine incorporation assays were performed to assess cell proliferation. Flow cytometry was conducted to detect alterations in the cell cycle and cell apoptosis. Western blotting and coimmunoprecipitation were performed to determine protein expression levels. In SCC15 cells, dioscin treatment significantly induced cell cycle arrest, increased apoptosis and inhibited proliferation compared with the control group. Mechanistically, the tumor suppressor protein Ras association domain­containing protein 1A (RASSF1A) was activated and oncoprotein yes­associated protein (YAP) was phosphorylated by dioscin. Furthermore, YAP overexpression and knockdown reduced and enhanced the inhibitory effects of dioscin on SCC15 cells, respectively. In summary, the results demonstrated that, compared with the control group, dioscin upregulated RASSF1A expression in OSCC cells, which resulted in YAP phosphorylation, thus weakening its transcriptional coactivation function, enhancing cell cycle arrest and apoptosis, and inhibiting cell proliferation. The present study indicated that dioscin may serve as a therapeutic agent for OSCC.


Asunto(s)
Antineoplásicos/farmacología , Carcinoma de Células Escamosas/metabolismo , Proliferación Celular/efectos de los fármacos , Diosgenina/análogos & derivados , Neoplasias de la Boca/metabolismo , Proteínas Supresoras de Tumor/metabolismo , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Apoptosis , Puntos de Control del Ciclo Celular , Línea Celular Tumoral , Diosgenina/farmacología , Humanos , Proteínas Serina-Treonina Quinasas/metabolismo , Factores de Transcripción/metabolismo , Proteínas Supresoras de Tumor/genética
20.
Nat Commun ; 12(1): 1703, 2021 03 17.
Artículo en Inglés | MEDLINE | ID: mdl-33731717

RESUMEN

The factors regulating cellular identity are critical for understanding the transition from health to disease and responses to therapies. Recent literature suggests that autophagy compromise may cause opposite effects in different contexts by either activating or inhibiting YAP/TAZ co-transcriptional regulators of the Hippo pathway via unrelated mechanisms. Here, we confirm that autophagy perturbation in different cell types can cause opposite responses in growth-promoting oncogenic YAP/TAZ transcriptional signalling. These apparently contradictory responses can be resolved by a feedback loop where autophagy negatively regulates the levels of α-catenins, LC3-interacting proteins that inhibit YAP/TAZ, which, in turn, positively regulate autophagy. High basal levels of α-catenins enable autophagy induction to positively regulate YAP/TAZ, while low α-catenins cause YAP/TAZ activation upon autophagy inhibition. These data reveal how feedback loops enable post-transcriptional determination of cell identity and how levels of a single intermediary protein can dictate the direction of response to external or internal perturbations.


Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Autofagia/fisiología , Transactivadores/metabolismo , Factores de Transcripción/metabolismo , alfa Catenina/metabolismo , Animales , Células Cultivadas , Células Epiteliales , Retroalimentación Fisiológica , Humanos , Ratones , Proteínas Asociadas a Microtúbulos/metabolismo , Mutación , Unión Proteica , Dominios y Motivos de Interacción de Proteínas/genética , Transducción de Señal , alfa Catenina/química , alfa Catenina/genética
SELECCIÓN DE REFERENCIAS
DETALLE DE LA BÚSQUEDA
...