RESUMEN
P38ß, p38γ, and p38δ have been sporadically and scarcely reported to be involved in the carcinogenesis of cancers, compared with p38α isoform. However, little has been known regarding their clinicopathological significance and biological roles in esophageal squamous cell carcinoma (ESCC). Expression status of p38ß, p38γ, and p38δ was assayed using immunohistochemistry with ESCC tissue microarray; ensuing clinicopathological significance was statistically analyzed. To define its biological roles on proliferation, migration and invasion of ESCC cell line Eca109 in vitro, MTT, wound healing, and Transwell assays were employed, respectively. As confirmation, athymic nude mice were taken to verify the effect over proliferation in vivo. It was found that both p38ß and p38δ expression, other than p38γ, were significantly higher in ESCC tissues compared with paired normal controls. In terms of prognosis, only p38ß expression was observed to be significantly associated with overall prognosis. Clinicopathologically, there was significant association between p38γ expression and clinical stage, lymph nodes metastases, and tumor volume. No significant association was found for p38ß and p38δ between its expression and other clinicopathological parameters other than significant difference of expression between ESCC versus normal control. In Eca109, it was observed that p38ß, p38γ, and p38δ can promote the cell growth and motility. As verification, over-expression of p38δ can promote, whereas knockdown of p38γ can prevent, the tumorigenesis in nude mice model xenografted with Eca109 cells whose basal level of p38δ was stably over-expressed and p38γ was stably knocked down. Together, our results demonstrate that p38ß, p38γ, and p38δ played oncogenic roles in ESCC.
Asunto(s)
Carcinoma de Células Escamosas/enzimología , Neoplasias Esofágicas/enzimología , Proteína Quinasa 11 Activada por Mitógenos/fisiología , Proteína Quinasa 12 Activada por Mitógenos/fisiología , Proteína Quinasa 13 Activada por Mitógenos/fisiología , Proteínas de Neoplasias/fisiología , Animales , Apoptosis , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patología , Ciclo Celular , Línea Celular Tumoral , Movimiento Celular , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/patología , Femenino , Xenoinjertos , Humanos , Metástasis Linfática , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Proteína Quinasa 11 Activada por Mitógenos/genética , Proteína Quinasa 12 Activada por Mitógenos/genética , Proteína Quinasa 13 Activada por Mitógenos/genética , Proteínas de Neoplasias/genética , Pronóstico , Interferencia de ARN , ARN Interferente Pequeño/genética , Proteínas Recombinantes de Fusión/metabolismo , Carga TumoralRESUMEN
Inhibition of p38MAPK alpha/beta is known to enhance 1,25-dihydroxyvitamin (1,25D)-induced monocytic differentiation, but the detailed mechanism of this effect was not clear. We now show that the enhancement of differentiation becomes apparent with slow kinetics (12-24 h). Interestingly, the inhibition of p38MAPK alpha/beta by their selective inhibitor SB202190 (SB) leads to an upregulated expression of p38MAPK isoforms gamma and delta in 1,25D-treated AML cells, in cell lines and in primary culture. Although the expression and activating phosphorylations of p38MAPK alpha are also increased by an exposure of the cells to SB, its kinase activity is blocked by SB, as shown by reduced levels of phosphorylated Hsp27, a downstream target of p38MAPK alpha. A positive role of p38MAPKs in 1,25D-induced differentiation is shown by the inhibition of differentiation by antisense oligonucleotides to all p38MAPK isoforms. Other principal branches of MAPK pathways showed early (6 h) activation of MEK/ERK by SB, followed by activation of JNK1/2 pathway and enhanced expression and/or activation of PU.1, ATF-2 differentiation-related transcription factors. Taken together with previous reports, the results indicate that 1,25D-induced differentiation is enhanced by the activation of at least three branches of MAPK pathways (ERK1/2; p38MAPK gamma/delta; JNK1/2). This activation may result from the removal of feedback inhibition of an upstream regulator of those pathways, when p38MAPK alpha and beta are inhibited by SB.
Asunto(s)
Calcitriol/farmacología , Diferenciación Celular/efectos de los fármacos , Leucemia Mieloide Aguda/patología , Proteína Quinasa 12 Activada por Mitógenos/fisiología , Proteína Quinasa 13 Activada por Mitógenos/fisiología , Antígenos de Diferenciación/genética , Antígenos de Diferenciación/metabolismo , Diferenciación Celular/genética , Relación Dosis-Respuesta a Droga , Regulación Enzimológica de la Expresión Génica/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Células HL-60 , Humanos , Imidazoles/farmacología , Isoenzimas/genética , Isoenzimas/metabolismo , Isoenzimas/fisiología , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Proteína Quinasa 12 Activada por Mitógenos/antagonistas & inhibidores , Proteína Quinasa 12 Activada por Mitógenos/genética , Proteína Quinasa 12 Activada por Mitógenos/metabolismo , Proteína Quinasa 13 Activada por Mitógenos/antagonistas & inhibidores , Proteína Quinasa 13 Activada por Mitógenos/genética , Proteína Quinasa 13 Activada por Mitógenos/metabolismo , Monocitos/efectos de los fármacos , Monocitos/metabolismo , Monocitos/fisiología , Inhibidores de Proteínas Quinasas/farmacología , Piridinas/farmacología , Factores de Tiempo , Células U937RESUMEN
The mitogen-activated protein kinase p38-gamma is highly expressed in skeletal muscle and is associated with the dystrophin glycoprotein complex; however, its function remains unclear. After induced damage, muscle in mice lacking p38-gamma generated significantly fewer myofibers than wild-type muscle. Notably, p38-gamma-deficient muscle contained 50% fewer satellite cells that exhibited premature Myogenin expression and markedly reduced proliferation. We determined that p38-gamma directly phosphorylated MyoD on Ser199 and Ser200, which results in enhanced occupancy of MyoD on the promoter of myogenin together with markedly decreased transcriptional activity. This repression is associated with extensive methylation of histone H3K9 together with recruitment of the KMT1A methyltransferase to the myogenin promoter. Notably, a MyoD S199A/S200A mutant exhibits markedly reduced binding to KMT1A. Therefore, p38-gamma signaling directly induces the assembly of a repressive MyoD transcriptional complex. Together, these results establish a hitherto unappreciated and essential role for p38-gamma signaling in positively regulating the expansion of transient amplifying myogenic precursor cells during muscle growth and regeneration.
Asunto(s)
Diferenciación Celular/genética , Silenciador del Gen , Proteína Quinasa 12 Activada por Mitógenos/fisiología , Desarrollo de Músculos/genética , Músculo Esquelético/citología , Animales , Línea Celular , Proliferación Celular , Epigénesis Genética , Regulación de la Expresión Génica , Histonas/metabolismo , Ratones , Ratones Endogámicos C57BL , Músculo Esquelético/metabolismo , Músculo Esquelético/fisiología , Proteína MioD/metabolismo , Miogenina/genética , Miogenina/metabolismo , Fosforilación , Regiones Promotoras Genéticas , Regeneración , Transducción de Señal , Transcripción GenéticaRESUMEN
We and others previously showed that p38 mitogen-activated protein kinase is indispensable for myogenic differentiation. However, it is less clear which of the four p38 isoforms in the mouse genome participates in this process. Using C2C12 myogenic cells as a model, we showed here that p38alpha, beta, and gamma are expressed with distinct expression patterns during differentiation. Knockdown of any of them by small interfering RNA inhibits myogenic differentiation, which suggests that the functions of the three p38 isoforms are not completely redundant. To further elucidate the unique role of each p38 isoform in myogenic differentiation, we individually knocked down one p38 isoform at a time in C2C12 cells, and we compared the whole-genome gene expression profiles by microarrays. We found that some genes are coregulated by all three p38 isoforms, whereas others are uniquely regulated by one particular p38 isoform. Furthermore, several novel p38 target genes (i.e., E2F2, cyclin D3, and WISP1) are found to be required for myogenin expression, which provides a molecular basis to explain why different p38 isoforms are required for myogenic differentiation.
Asunto(s)
Proteína Quinasa 11 Activada por Mitógenos/fisiología , Proteína Quinasa 12 Activada por Mitógenos/fisiología , Proteína Quinasa 14 Activada por Mitógenos/fisiología , Desarrollo de Músculos/fisiología , Animales , Secuencia de Bases , Proteínas CCN de Señalización Intercelular , Diferenciación Celular , Línea Celular , Ciclina D3 , Ciclinas/genética , Ciclinas/metabolismo , Factor de Transcripción E2F2/genética , Factor de Transcripción E2F2/metabolismo , Perfilación de la Expresión Génica , Ratones , Proteína Quinasa 11 Activada por Mitógenos/antagonistas & inhibidores , Proteína Quinasa 11 Activada por Mitógenos/genética , Proteína Quinasa 12 Activada por Mitógenos/antagonistas & inhibidores , Proteína Quinasa 12 Activada por Mitógenos/genética , Proteína Quinasa 14 Activada por Mitógenos/antagonistas & inhibidores , Proteína Quinasa 14 Activada por Mitógenos/genética , Modelos Biológicos , Desarrollo de Músculos/genética , Músculos/enzimología , Mioblastos/citología , Mioblastos/enzimología , Análisis de Secuencia por Matrices de Oligonucleótidos , Proteínas Oncogénicas/genética , Proteínas Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas , ARN Mensajero/genética , ARN Mensajero/metabolismo , ARN Interferente Pequeño/genéticaRESUMEN
The regulation of skeletal muscle formation (myogenesis) is essential for normal development as well as in pathological conditions such as muscular dystrophies and inflammatory myopathies. Findings published over the past years have established a key role for the p38 MAP kinase signaling pathway in the control of muscle gene expression and myotube formation. However, the relative contribution of the four p38 MAP kinases (p38alpha, p38beta, p38gamma and p38delta) to this process was unknown. We have recently demonstrated that myoblasts lacking p38alpha, but not those lacking p38beta or p38delta, were unable to differentiate and form multinucleated myotubes, while p38gamma-deficient myoblasts exhibited an attenuated fusion capacity. Defective myogenesis in the absence of p38alpha was attributed to delayed cell cycle exit and continuous proliferation in differentiation-promoting conditions, caused by enhanced activation of the JNK/cJun pathway. We discuss these findings in the context of the emerging crosstalk of p38 and JNK signaling pathways in controlling cell growth and differentiation.
Asunto(s)
Ciclo Celular/fisiología , Proteínas Quinasas JNK Activadas por Mitógenos/fisiología , Proteína Quinasa 14 Activada por Mitógenos/fisiología , Mioblastos/citología , Animales , Animales Recién Nacidos , Proteínas de Ciclo Celular/biosíntesis , Proteínas de Ciclo Celular/genética , Diferenciación Celular/fisiología , División Celular/fisiología , Fusión Celular , Células Cultivadas/citología , Activación Enzimática , Regulación de la Expresión Génica , Humanos , Ratones , Ratones Noqueados , Proteína Quinasa 11 Activada por Mitógenos/fisiología , Proteína Quinasa 12 Activada por Mitógenos/fisiología , Proteína Quinasa 13 Activada por Mitógenos/fisiología , Proteína Quinasa 14 Activada por Mitógenos/deficiencia , Proteína Quinasa 14 Activada por Mitógenos/genética , Fibras Musculares Esqueléticas/citología , Proteínas Musculares/biosíntesis , Proteínas Musculares/genética , Proteínas de Neoplasias/metabolismo , Especificidad de Órganos , Fosforilación , Procesamiento Proteico-Postraduccional , Rabdomiosarcoma/patologíaRESUMEN
Activation of the p38 MAP kinase pathways is crucial for the adaptation of mammalian cells to changes in the osmolarity of the environment. Here we identify SAP97/hDlg, the mammalian homologue of the Drosophila tumour suppressor Dlg, as a physiological substrate for the p38gamma MAP kinase (SAPK3/p38gamma) isoform. SAP97/hDlg is a scaffold protein that forms multiprotein complexes with a variety of proteins and is targeted to the cytoskeleton by its association with the protein guanylate kinase-associated protein (GKAP). The SAPK3/p38gamma-catalysed phosphorylation of SAP97/hDlg triggers its dissociation from GKAP and therefore releases it from the cytoskeleton. This is likely to regulate the integrity of intercellular-junctional complexes, and cell shape and volume in response to osmotic stress.
Asunto(s)
Citoesqueleto/metabolismo , Proteína Quinasa 12 Activada por Mitógenos/fisiología , Proteínas del Tejido Nervioso/metabolismo , Proteínas Adaptadoras Transductoras de Señales , Animales , Línea Celular Tumoral , Forma de la Célula/fisiología , Tamaño de la Célula , Citoesqueleto/química , Homólogo 1 de la Proteína Discs Large , Proteínas de Drosophila , Guanilato-Quinasas , Humanos , Uniones Intercelulares/metabolismo , Proteínas de la Membrana , Ratones , Ratones Noqueados , Proteína Quinasa 12 Activada por Mitógenos/genética , Proteínas del Tejido Nervioso/análisis , Presión Osmótica , Fosforilación , Proteínas Asociadas a SAP90-PSD95 , Especificidad por SustratoRESUMEN
Signal transduction pathways in eukaryotic cells integrate diverse extracellular signals, and regulate complex biological responses such as growth, differentiation and death. One group of proline-directed Ser/Thr protein kinases, the mitogen-activated protein kinases (MAPKs), plays a central role in these signalling pathways. Much attention has focused in recent years on three subfamilies of MAPKs, the extracellular signal regulated kinases (ERKs), c-Jun N-terminal kinases (JNKs) and the p38 MAPKs. However, the ERK family is broader than the ERK1 and ERK2 proteins that have been the subject of most studies in this area. Here we overview the work on ERKs 3 to 8, emphasising where possible their biological activities as well as distinctive biochemical properties. It is clear from these studies that these additional ERKs show similarities to ERK1 and ERK2, but with some interesting differences that challenge the paradigm of the archetypical ERK1/2 MAPK pathway.