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The objective of this Special Issue was to collate recent advances in the understanding of MAPKs' functions, particularly their roles in various pathologies, which constitute one of the most dynamic areas in cell signaling research [...].
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Proteínas Quinasas Activadas por Mitógenos , Humanos , Animales , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Proteínas Quinasas Activadas por Mitógenos/genética , Sistema de Señalización de MAP Quinasas , Transducción de SeñalRESUMEN
The microbiota in humans and animals play crucial roles in defense against pathogens and offer a promising natural source for immunomodulatory products. However, the development of physiologically relevant model systems and protocols for testing such products remains challenging. In this study, we present an experimental condition where various natural products derived from the registered lactic acid bacteria Ligilactobacillus salivarius CECT 9609, known for their immunomodulatory activity, were tested. These products included live and inactivated bacteria, as well as fermentation products at different concentrations and culture times. Using our established model system, we observed no morphological changes in the airway epithelium upon exposure to Pasteurella multocida, a common respiratory pathogen. However, early molecular changes associated with the innate immune response were detected through transcript analysis. By employing diverse methodologies ranging from microscopy to next-generation sequencing (NGS), we characterized the interaction of these natural products with the airway epithelium and their potential beneficial effects in the presence of P. multocida infection. In particular, our discovery highlights that among all Ligilactobacillus salivarius CECT 9609 products tested, only inactivated cells preserve the conformation and morphology of respiratory epithelial cells, while also reversing or altering the natural immune responses triggered by Pasteurella multocida. These findings lay the groundwork for further exploration into the protective role of these bacteria and their derivatives.
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Productos Biológicos , Ligilactobacillus salivarius , Infecciones por Pasteurella , Pasteurella multocida , Humanos , Animales , Inmunidad Innata , Células Epiteliales , Productos Biológicos/farmacología , Infecciones por Pasteurella/microbiología , Infecciones por Pasteurella/veterinariaRESUMEN
Proper physiological function of mammalian airways requires the differentiation of basal stem cells into secretory or multiciliated cells, among others. In addition, the self-renewal ability of these basal stem cells is crucial for developing a quick response to toxic agents in order to re-establish the epithelial barrier function of the airways. Although these epithelial missions are vital, little is known about those mechanism controlling airway epithelial regeneration in health and disease. p53 has been recently proposed as the guardian of homeostasis, promoting differentiation programs, and antagonizing a de-differentiation program. Here, we exploit mouse and human tracheal epithelial cell culture models to study the role of MDM2-p53 signaling in self-renewal and differentiation in the airway epithelium. We show that p53 protein regulation by MDM2 is crucial for basal stem cell differentiation and to keep proper cell proliferation. Therefore, we suggest that MDM2/p53 interaction modulation is a potential target to control regeneration of the mammalian airway epithelia without massively affecting the epithelium integrity and differentiation potential.
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Diferenciación Celular/fisiología , Epitelio/metabolismo , Proteínas Proto-Oncogénicas c-mdm2/metabolismo , Mucosa Respiratoria/metabolismo , Células Madre/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Animales , Proliferación Celular/fisiología , Células Epiteliales/metabolismo , Femenino , Homeostasis/fisiología , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Regeneración/fisiología , Transducción de Señal/fisiología , Tráquea/metabolismoRESUMEN
Adipose tissue has emerged as an important regulator of whole-body metabolism, and its capacity to dissipate energy in the form of heat has acquired a special relevance in recent years as potential treatment for obesity. In this context, the p38MAPK pathway has arisen as a key player in the thermogenic program because it is required for the activation of brown adipose tissue (BAT) thermogenesis and participates also in the transformation of white adipose tissue (WAT) into BAT-like depot called beige/brite tissue. Here, using mice that are deficient in p38α specifically in adipose tissue (p38αFab-KO), we unexpectedly found that lack of p38α protected against high-fat diet (HFD)-induced obesity. We also showed that p38αFab-KO mice presented higher energy expenditure due to increased BAT thermogenesis. Mechanistically, we found that lack of p38α resulted in the activation of the related protein kinase family member p38δ. Our results showed that p38δ is activated in BAT by cold exposure, and lack of this kinase specifically in adipose tissue (p38δ Fab-KO) resulted in overweight together with reduced energy expenditure and lower body and skin surface temperature in the BAT region. These observations indicate that p38α probably blocks BAT thermogenesis through p38δ inhibition. Consistent with the results obtained in animals, p38α was reduced in visceral and subcutaneous adipose tissue of subjects with obesity and was inversely correlated with body mass index (BMI). Altogether, we have elucidated a mechanism implicated in physiological BAT activation that has potential clinical implications for the treatment of obesity and related diseases such as diabetes.
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Tejido Adiposo Pardo/enzimología , Tejido Adiposo Pardo/fisiología , Proteína Quinasa 13 Activada por Mitógenos/antagonistas & inhibidores , Proteína Quinasa 14 Activada por Mitógenos/metabolismo , Termogénesis , Adipocitos Marrones/enzimología , Adulto , Animales , Índice de Masa Corporal , Diabetes Mellitus Experimental/enzimología , Diabetes Mellitus Experimental/prevención & control , Dieta , Metabolismo Energético , Activación Enzimática , Humanos , Sistema de Señalización de MAP Quinasas , Masculino , Ratones Endogámicos C57BL , Ratones Noqueados , Proteína Quinasa 13 Activada por Mitógenos/metabolismo , Modelos Biológicos , Obesidad/enzimología , Obesidad/prevención & control , Proteína Desacopladora 1/metabolismoRESUMEN
The feasibility of obtaining resistant starch type III (RS3) from malanga flour (Xanthosoma sagittifolium), as an unconventional source of starch, was evaluated using the hydrothermal treatment of autoclaving. The physicochemical characterization of RS3 made from malanga flour was carried out through the evaluation of the chemical composition, color attributes, and thermal properties. In addition, the contents of the total starch, available starch, resistant starch, and retrograded resistant starch were determined by in vitro enzymatic tests. A commercial corn starch sample was used to produce RS3 and utilized to compare all of the analyses. The results showed that native malanga flour behaved differently in most of the evaluations performed, compared to the commercial corn starch. These results could be explained by the presence of minor components that could interfere with the physicochemical and functional properties of the flour; however, the RS3 samples obtained from malanga flour and corn starch were similar in their thermal and morphological features, which may be related to their similarities in the content and molecular weight of amylose, in both of the samples. Furthermore, the yields for obtaining the autoclaved powders from corn starch and malanga flour were similar (≈89%), which showed that the malanga flour is an attractive raw material for obtaining RS3 with adequate yields, to be considered in the subsequent research.
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Harina , Almidón Resistente , Xanthosoma/química , Zea mays/químicaRESUMEN
High molecular weight chitosan (≈322 kDa) was obtained from chitin isolated from Brachystola magna (Girard) to produced biodegradable films. Their physicochemical, mechanical and water vapor permeability (WVP) properties were compared against commercial chitosan films with different molecular weights. Brachystola magna chitosan films (CFBM) exhibited similar physicochemical and mechanical characteristics to those of commercial chitosans. The CFBM films presented lower WVP values (10.01 × 10-11 g/m s Pa) than commercial chitosans films (from 16.06 × 10-11 to 64.30 × 10-11 g/m s Pa). Frankfurt-type sausages were covered with chitosan films and stored in refrigerated conditions (4 °C). Their quality attributes (color, weight loss, pH, moisture, texture and lipid oxidation) were evaluated at 0, 5, 10, 15 and 20 days. Sausages covered with CFMB films presented the lowest weight loss (from 1.24% to 2.38%). A higher increase in hardness (from 22.32 N to 30.63 N) was observed in sausages covered with CFMB films. Compared with other films and the control (uncovered sausages), CFMB films delay pH reduction. Moreover, this film presents the lower lipid oxidation level (0.10 malonaldehyde mg/sample kg). Thus, chitosan of B. magna could be a good alternative as packaging material for meat products with high-fat content.
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Quitosano/química , Embalaje de Alimentos , Conservación de Alimentos , Saltamontes/química , Productos de la Carne , Membranas Artificiales , AnimalesRESUMEN
Growing evidence suggests that Diabetes Mellitus increases the risk of developing Alzheimer's disease. It is well known that hyperglycemia, a key feature of Diabetes Mellitus, may induce plasma osmolarity disturbances. Both hyperglycemia and hyperosmolarity promote the altered post-translational regulation of microtubule-associated protein Tau. Interestingly, abnormal hyperphosphorylation and cleavage of Tau have been proven to lead to the genesis of filamentous structures referred to as neurofibrillary tangles, the main pathological hallmark of Alzheimer's disease. We have previously described that hyperosmotic stress induced by sorbitol promotes Tau proteolysis and apoptosis in SH-SY5Y cells via caspase-3 activation. In order to gain insights into the regulatory mechanisms of such processes, in this work we explored the intracellular signaling pathways that regulate these events. We found that sorbitol treatment significantly enhanced the activation of conventional families of MAPK in SH-SY5Y cells. Tau proteolysis was completely prevented by JNK inhibition but not affected by either ERK1/2 or p38 MAPK blockade. Moreover, inhibition of JNK, but not ERK1/2 or p38 MAPK, efficiently prevented sorbitol-induced apoptosis and caspase-3 activation. In summary, we provide evidence that JNK signaling pathway is an upstream regulator of hyperosmotic stress-induced Tau cleavage and apoptosis in SH-SY5Y through the control of caspase-3 activation.
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Apoptosis/efectos de los fármacos , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Presión Osmótica/efectos de los fármacos , Proteolisis/efectos de los fármacos , Sorbitol/farmacología , Proteínas tau/metabolismo , Apoptosis/genética , Caspasa 3/genética , Caspasa 3/metabolismo , Línea Celular Tumoral , Activación Enzimática/efectos de los fármacos , Activación Enzimática/genética , Quinasas MAP Reguladas por Señal Extracelular/genética , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Humanos , Sistema de Señalización de MAP Quinasas/genética , Proteínas tau/genéticaRESUMEN
Tau is a microtubule-associated protein implicated in the pathogenesis of Alzheimer's disease and other related tauopathies. In this subset of neurodegenerative disorders, Tau auto-assembles into insoluble fibrils that accumulate in neurons as paired helical filaments (PHFs), promoting cellular dysfunction and cytotoxic effects. Growing evidence suggests that abnormal post-translational regulation, mainly hyperphosphorylation and aberrant cleavage, drives Tau to this pathological state. In this work we show that sorbitol-induced hyperosmotic stress promotes Tau proteolysis in SH-SY5Y neuroblastoma cells. The appearance of cleaved Tau was preceded by the activation of µ-calpain, the proteasome system and caspase-3. Tau proteolysis was completely prevented by caspase-3 inhibition but unaffected by neither the proteasome system nor µ-calpain activity blockade. Concomitantly, hyperosmotic stress induced apoptosis in SH-SY5Y cells, which was efficiently avoided by the inhibition of caspase-3 activity. Altogether, our results provide the first evidence that Tau protein is susceptible to caspase-3 proteolysis under hyperosmotic stress and suggest a positive relationship between Tau proteolysis and apoptosis in SH-SY5Y cells. J. Cell. Biochem. 117: 2781-2790, 2016. © 2016 Wiley Periodicals, Inc.
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Apoptosis , Caspasa 3/metabolismo , Regulación Neoplásica de la Expresión Génica , Neuroblastoma/patología , Presión Osmótica , Complejo de la Endopetidasa Proteasomal/metabolismo , Proteínas tau/metabolismo , Western Blotting , Proliferación Celular , Activación Enzimática , Humanos , Neuroblastoma/metabolismo , Fosforilación , Proteolisis , Células Tumorales CultivadasRESUMEN
Activating transcription factor 4 (Atf4), which is modulated by the protein kinase RNA-like ER kinase (PERK), is a stress-induced transcription factor responsible for controlling the expression of a wide range of adaptive genes, enabling cells to withstand stressful conditions. However, the impact of the Atf4 signaling pathway on airway regeneration remains poorly understood. In this study, we used mouse airway epithelial cell culture models to investigate the role of PERK/Atf4 in respiratory tract differentiation. Through pharmacological inhibition and silencing of ATF4, we uncovered the crucial involvement of PERK/Atf4 in the differentiation of basal stem cells, leading to a reduction in the number of secretory cells. ChIP-seq analysis revealed direct binding of ATF4 to regulatory elements of genes associated with osteoblast differentiation and secretory cell function. Our findings provide valuable insights into the role of ATF4 in airway epithelial differentiation and its potential involvement in innate immune responses and cellular adaptation to stress.
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Estrés del Retículo Endoplásmico , eIF-2 Quinasa , Animales , Ratones , eIF-2 Quinasa/genética , Estrés del Retículo Endoplásmico/genética , Factor de Transcripción Activador 4/genética , Factor de Transcripción Activador 4/metabolismo , Diferenciación Celular/genética , Sistema Respiratorio/metabolismoRESUMEN
Activation of p38γ modulates the integrity of the complex formed by the human discs large protein (hDlg) with cytoskeletal proteins, which is important for cell adaptation to changes in environmental osmolarity. Here we report that, in response to hyperosmotic stress, p38γ also regulates formation of complexes between hDlg and the nuclear protein polypyrimidine tract-binding protein-associated-splicing factor (PSF). Following osmotic shock, p38γ in the cell nucleus increases its association with nuclear hDlg, thereby causing dissociation of hDlg-PSF complexes. Moreover, hDlg and PSF bind different RNAs; in response to osmotic shock, p38γ causes hDlg-PSF and hDlg-RNA dissociation independently of its kinase activity. These findings identify a novel nuclear complex and suggest a previously unreported function of p38γ, which is independent of its catalytic activity and could affect mRNA processing and/or gene transcription to aid cell adaptation to osmolarity changes in the environment.
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Proteínas Adaptadoras Transductoras de Señales/metabolismo , Proteínas de la Membrana/metabolismo , Proteína Quinasa 12 Activada por Mitógenos/metabolismo , Presión Osmótica/fisiología , Proteínas de Unión al ARN/metabolismo , ARN/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Animales , Línea Celular , Homólogo 1 de la Proteína Discs Large , Células HeLa , Humanos , Inmunoprecipitación , Proteínas de la Membrana/genética , Ratones , Ratones Mutantes , Proteína Quinasa 12 Activada por Mitógenos/genética , Factor de Empalme Asociado a PTB , Fosforilación/genética , Fosforilación/fisiología , Reacción en Cadena de la Polimerasa , Unión Proteica/genética , Unión Proteica/fisiología , Proteínas de Unión al ARN/genética , Espectrometría de Masa por Láser de Matriz Asistida de Ionización DesorciónRESUMEN
Yearly, 1,500,000 cases of leishmaniasis are diagnosed, causing thousands of deaths. To advance in its therapy, we present an interdisciplinary protocol that unifies ethnobotanical knowledge of natural compounds and the latest bioinformatics advances to respond to an orphan disease such as leishmaniasis and specifically the one caused by Leishmania amazonensis. The use of ethnobotanical information serves as a basis for the development of new drugs, a field in which computer-aided drug design (CADD) has been a revolution. Taking this information from Amazonian communities, located in the area with a high prevalence of this disease, a protocol has been designed to verify new leads. Moreover, a method has been developed that allows the evaluation of lead molecules, and the improvement of their affinity and specificity against therapeutic targets. Through this approach, deguelin has been identified as a good lead to treat the infection due to its potential as an ornithine decarboxylase (ODC) inhibitor, a key enzyme in Leishmania development. Using an in silico-generated combinatorial library followed by docking approaches, we have found deguelin derivatives with better affinity and specificity against ODC than the original compound, suggesting that this approach could be adapted for developing new drugs against leishmaniasis.
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Tight-junction (TJ) proteins are essential for establishing the barrier function between neighbor epithelial cells, but also for recognition of pathogens or cell migration. Establishing the expression pattern and localization of different TJ proteins will help to understand the development and physiology of the airway. Here we identify that the junctional adhesion molecule 3 (Jam3) expression is restricted to multiciliated cells (MCCs) in the airway epithelium. In vitro, Jam3 expression varies along airway basal stem cell (BSC) differentiation and upon DAPT treatment or IL6 exposure. However, Jam3 is not required for BSC differentiation to specific cell types. In addition, we found that MCC lacking Jam3 display normal cilia morphology and cilia beating frequency with a delay in BB assembly/positioning in MCCs during differentiation. Remarkably, Jam3 in MCC is mostly localized to subapical organelles, which are negative for the apical recycling endosome marker Rab11 and positive for EEA1. Our data show that Jam3 expression is connected to mature MCC in the airway epithelium and suggest a Jam3 role unrelated to its known barrier function.
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IL6 is an essential cytokine in metabolism regulation and for intercommunication among different organs and tissues. IL6 produced by different tissues has different functions and therefore it is very important to understand the mechanism of its expression in adipose tissue. In this work we demonstrated that IL6 expression in mouse preadipocytes, like in human, is partially dependent on Wnt5a and JNK. Using mouse preadipocytes lacking each one of the p38 SAPK family members, we have shown that IL6 expression is also p38γ and p38δ dependent. In fact, the lack of some of these two kinases increases IL6 expression without altering that of Wnt5a. Moreover, we show that the absence of p38δ promotes greater ERK1/2 phosphorylation in a MEK1/2 independent manner, and that this increased ERK1/2 phosphorylation state is contributing to the higher IL6 expression in p38δ-/- preadipocytes. These results suggest a new crosstalk between two MAPK signaling pathway, p38δ and ERK1/2, where p38δ modulates the phosphorylation state of ERK1/2.
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This paper is focused on demonstrating with a real case that Ethnobotany added to Bioinformatics is a promising tool for new drugs search. It encourages the in silico investigation of "challua kaspi", a medicinal kichwa Amazonian plant (Aspidosperma spruceanum) against a Neglected Tropical Disease, leishmaniasis. The illness affects over 150 million people especially in subtropical regions, there is no vaccination and conventional treatments are unsatisfactory. In attempts to find potent and safe inhibitors of its etiological agent, Leishmania, we recovered the published traditional knowledge on kichwa antimalarials and selected three A. spruceanum alkaloids, (aspidoalbine, aspidocarpine and tubotaiwine), to evaluate by molecular docking their activity upon five Leishmania targets: DHFR-TS, PTR1, PK, HGPRT and SQS enzymes. Our simulation results suggest that aspidoalbine interacts competitively with the five targets, with a greater affinity for the active site of PTR1 than some physiological ligands. Our virtual data also point to the demonstration of few side effects. The predicted binding free energy has a greater affinity to Leishmania proteins than to their homologous in humans (TS, DHR, PKLR, HGPRT and SQS), and there is no match with binding pockets of physiological importance. Keys for the in silico protocols applied are included in order to offer a standardized method replicable in other cases. Apocynaceae having ethnobotanical use can be virtually tested as molecular antileishmaniasis new drugs.
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Organelle positioning as many other morphological parameters in a cell is not random. Centriole positioning as centrosomes or ciliary basal bodies is not an exception to this rule in cell biology. Indeed, centriole positioning is a tightly regulated process that occurs during development, and it is critical for many organs to function properly, not just during development but also in the adulthood. In this book chapter, we overview our knowledge on centriole positioning in different and highly specialized animal cells like photoreceptor or ependymal cells. We will also discuss recent advances in the discovery of molecular pathways involved in this process, mostly related to the cytoskeleton and the cell polarity pathways. And finally, we present quantitative methods that have been used to assess centriole positioning in different cell types although mostly in epithelial cells.
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Compartimento Celular , Centriolos/metabolismo , Animales , Polaridad Celular , Citoesqueleto/metabolismoRESUMEN
Increasing the thermogenic capacity of adipose tissue to enhance organismal energy expenditure is considered a promising therapeutic strategy to combat obesity. Here, we report that expression of the p38 MAPK activator MKK6 is elevated in white adipose tissue of obese individuals. Using knockout animals and shRNA, we show that Mkk6 deletion increases energy expenditure and thermogenic capacity of white adipose tissue, protecting mice against diet-induced obesity and the development of diabetes. Deletion of Mkk6 increases T3-stimulated UCP1 expression in adipocytes, thereby increasing their thermogenic capacity. Mechanistically, we demonstrate that, in white adipose tissue, p38 is activated by an alternative pathway involving AMPK, TAK, and TAB. Our results identify MKK6 in adipocytes as a potential therapeutic target to reduce obesity.Brown and beige adipose tissues dissipate heat via uncoupling protein 1 (UCP1). Here the authors show that the stress activated kinase MKK6 acts as a repressor of UCP1 expression, suggesting that its inhibition promotes adipose tissue browning and increases organismal energy expenditure.
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Tejido Adiposo Blanco/enzimología , MAP Quinasa Quinasa 6/metabolismo , Obesidad/enzimología , Proteína Desacopladora 1/metabolismo , Adipocitos Blancos/metabolismo , Adulto , Anciano , Animales , Estudios de Casos y Controles , Diabetes Mellitus/etiología , Dieta Alta en Grasa , Metabolismo Energético , Femenino , Humanos , Sistema de Señalización de MAP Quinasas , Masculino , Síndrome Metabólico/etiología , Ratones Endogámicos C57BL , Ratones Noqueados , Persona de Mediana Edad , Obesidad/etiología , Triyodotironina/fisiología , Núcleo Hipotalámico Ventromedial/metabolismoRESUMEN
The biochemical mechanism of apoptosis induced by ceramide remains still unclear, although it has been reported that dephosphorylation of PKB at Ser-473 may be a key event. In this article, we show that C(2)-ceramide (N-acetyl-sphingosine) induces the dephosphorylation of both protein kinase B (PKB) and glycogen synthase kinase-3 (GSK3) in cerebellar granule cells (CGC). We also show that lithium protects against the apoptosis induced by C(2)-ceramide by blocking the dephosphorylation of both kinases. Since lithium inhibits in vivo the observed protein phosphatase-2A (PP2A) activation induced by ceramide, we hypothesise that the neuroprotective action of lithium may be due to the inhibition of the PP2A activation by apoptotic stimuli.
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Apoptosis/efectos de los fármacos , Cerebelo/enzimología , Litio/farmacología , Fosfoproteínas Fosfatasas/metabolismo , Proteínas Serina-Treonina Quinasas , Esfingosina/análogos & derivados , Esfingosina/antagonistas & inhibidores , Animales , Proteínas Quinasas Dependientes de Calcio-Calmodulina/metabolismo , Células Cultivadas , Cerebelo/citología , Cerebelo/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Inhibidores Enzimáticos/farmacología , Glucógeno Sintasa Quinasa 3 , Glucógeno Sintasa Quinasas , Fármacos Neuroprotectores/farmacología , Fosforilación , Proteína Fosfatasa 2 , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas c-akt , Ratas , Ratas WistarRESUMEN
SAPK3 (stress-activated protein kinase-3, also known as p38gamma) is a member of the mitogen-activated protein kinase family; it phosphorylates substrates in response to cellular stress, and has been shown to bind through its C-terminal sequence to the PDZ domain of alpha1-syntrophin. In the present study, we show that SAP90 [(synapse-associated protein 90; also known as PSD-95 (postsynaptic density-95)] is a novel physiological substrate for both SAPK3/p38gamma and the ERK (extracellular-signal-regulated protein kinase). SAPK3/p38gamma binds preferentially to the third PDZ domain of SAP90 and phosphorylates residues Thr287 and Ser290 in vitro, and Ser290 in cells in response to cellular stresses. Phosphorylation of SAP90 is dependent on the binding of SAPK3/p38gamma to the PDZ domain of SAP90. It is not blocked by SB 203580, which inhibits SAPK2a/p38alpha and SAPK2b/p38beta but not SAPK3/p38gamma, or by the ERK pathway inhibitor PD 184352. However, phosphorylation is abolished when cells are treated with a cell-permeant Tat fusion peptide that disrupts the interaction of SAPK3/p38gamma with SAP90. ERK2 also phosphorylates SAP90 at Thr287 and Ser290 in vitro, but this does not require PDZ-dependent binding. SAP90 also becomes phosphorylated in response to mitogens, and this phosphorylation is prevented by pretreatment of the cells with PD 184352, but not with SB 203580. In neurons, SAP90 and SAPK3/p38gamma co-localize and they are co-immunoprecipitated from brain synaptic junctional preparations. These results demonstrate that SAP90 is a novel binding partner for SAPK3/p38gamma, a first physiological substrate described for SAPK3/p38gamma and a novel substrate for ERK1/ERK2, and that phosphorylation of SAP90 may play a role in regulating protein-protein interactions at the synapse in response to adverse stress- or mitogen-related stimuli.
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Sistema de Señalización de MAP Quinasas/fisiología , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Mitógenos/farmacología , Proteínas del Tejido Nervioso/metabolismo , Procesamiento Proteico-Postraduccional/efectos de los fármacos , Estrés Fisiológico/metabolismo , Secuencia de Aminoácidos , Animales , Benzamidas/farmacología , Línea Celular/metabolismo , Línea Celular/efectos de la radiación , Activación Enzimática/efectos de la radiación , Inhibidores Enzimáticos/farmacología , Humanos , Imidazoles/farmacología , Ratones , Microscopía Fluorescente , Proteína Quinasa 3 Activada por Mitógenos , Datos de Secuencia Molecular , Proteínas del Tejido Nervioso/química , Neuronas/metabolismo , Neuronas/ultraestructura , Presión Osmótica , Células PC12/metabolismo , Células PC12/efectos de la radiación , Fosforilación/efectos de los fármacos , Fosfoserina/análisis , Fosfotreonina/análisis , Mapeo de Interacción de Proteínas , Estructura Terciaria de Proteína , Piridinas/farmacología , Ratas , Ratas Sprague-Dawley , Proteínas Recombinantes de Fusión/metabolismo , Proteínas Asociadas a SAP90-PSD95 , Rayos Ultravioleta , Proteínas Quinasas p38 Activadas por MitógenosRESUMEN
We investigated apoptosis induced by the green tea component the epigallocatechin-3-gallate (EGCG) and the pathways underlying its activity in a colon cancer cell line. A complete understanding of the mechanism(s) and molecules targeted by green tea polyphenols could be useful in developing novel therapeutic approaches for cancer treatment. EGCG, which is the major polyphenol in green tea, has cytotoxic effects and induced cell death in HT-29 cell death. In this study, we evaluated the effect EGCG on mitogen-activated protein kinase (MAPK) and Akt pathways. EGCG treatment increased phospho-ERK1/2, -JNK1/2 and -p38α, -p38γ and -p38δ, as well as phospho-Akt levels. Using a combination of kinase inhibitors, we found that EGCG-induced cell death is partially blocked by inhibiting Akt, ERK1/2 or alternative p38MAPK activity. Our data suggest that these kinase pathways are involved in the anti-cancer effects of EGCG and indicate potential use of this compound as chemotherapeutic agent for colon cancer treatment.
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Anticarcinógenos/metabolismo , Antioxidantes/metabolismo , Apoptosis , Catequina/análogos & derivados , Neoplasias del Colon/metabolismo , Sistema de Señalización de MAP Quinasas , Proteínas Proto-Oncogénicas c-akt/agonistas , Apoptosis/efectos de los fármacos , Catequina/metabolismo , Línea Celular Tumoral , Neoplasias del Colon/inducido químicamente , Neoplasias del Colon/enzimología , Neoplasias del Colon/prevención & control , Manipulación de Alimentos , Células HEK293 , Humanos , Isoenzimas/antagonistas & inhibidores , Isoenzimas/química , Isoenzimas/genética , Isoenzimas/metabolismo , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Concentración Osmolar , Fosforilación/efectos de los fármacos , Inhibidores de Proteínas Quinasas/farmacología , Procesamiento Proteico-Postraduccional/efectos de los fármacos , Proteínas Proto-Oncogénicas c-akt/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-akt/metabolismo , Interferencia de ARN , ARN Interferente Pequeño , Té/química , Proteínas Quinasas p38 Activadas por Mitógenos/antagonistas & inhibidores , Proteínas Quinasas p38 Activadas por Mitógenos/química , Proteínas Quinasas p38 Activadas por Mitógenos/genética , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
Human disc-large (hDlg) is a scaffold protein critical for the maintenance of cell polarity and adhesion. hDlg is a component of the p38gamma MAP kinase pathway, which is important for the adaptation of mammalian cells to changes in environmental osmolarity. Here we report a strong decrease in the levels of hDlg protein in the human epithelial cell line HeLa when exposed to osmotic shock. This is independent of the phosphorylation state of hDlg, is prevented by preincubating the cell with the caspase inhibitor z-VAD and is part of the apoptotic process triggered by cellular stress. Although, both caspase 3 and caspase 6 are strongly activated by osmotic shock, the time course of caspase 6 activation parallels hDlg degradation, suggesting that this caspase may be responsible for the proteolysis. Mutating hDlg Asp747 to Ala abolishes caspase-induced cleavage, but does not affect the early stage of apoptosis or cell attachment. Our findings show that osmotic stress triggers hDlg degradation through a mechanism different from the one mediated by proteasomes, and we identify hDlg as a caspase substrate during the apoptotic process, although its proteolysis may not be implicated in the progression of early apoptosis.