RESUMEN
The ras oncogene needs a second factor to induce transformation and tumorigenicity of the cell. In this study, we show that mouse fibroblast 7-4-Stat3C cells overexpressing both Ha-ras(val12) oncogene and active-form Stat3 (Stat3C) showed higher colony formation in soft agar and xenograft tumor growth in BALB/c mice. Further studies show that both serine-727 and tyrosine-705 of Stat3 were phosphorylated while Ha-ras was overexpressed. Interleukin-6 (IL-6)-induced phosphorylation of tyrosine-705 and serine-727, as well as DNA-binding and transcriptional activity of Stat3 were further enhanced by Ha-ras overexpression. In addition, overexpression of Stat3C in 7-4-Stat3C cells prevented the cells from morphological change and apoptosis triggered by the Ha-ras oncogene under serum-depleted conditions. We demonstrate that Ha-ras and Stat3 acting together synergistically induce Stat3 phosphorylation at serine-727 phosphorylation and cyclin D1 expression and further enhance transformation and tumorigenicity of the cell. Ha-ras-induced Stat3 phosphorylation at serine-727 plays a pivotal role in transcriptional activation of cyclin D1 and suppression of cell apoptosis. The effect of Ha-ras on Stat3 phosphorylation at serine-727 was also detected in human bladder (T24) and lung (H460) cancer cells. Stat3 phosphorylation at serine-727 is important in Ras-related tumorigenesis.
Asunto(s)
Transformación Celular Neoplásica , Ciclina D1/genética , Genes ras , Interleucina-6/farmacología , Fosforilación/efectos de los fármacos , Factor de Transcripción STAT3/genética , Serina/metabolismo , Secuencia de Aminoácidos , Animales , Apoptosis , Línea Celular Tumoral , Transformación Celular Neoplásica/genética , Transformación Celular Neoplásica/metabolismo , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patología , Ratones , Trasplante de Neoplasias , Factor de Transcripción STAT3/metabolismo , Activación Transcripcional/genética , Regulación hacia Arriba/genética , Neoplasias de la Vejiga Urinaria/metabolismo , Neoplasias de la Vejiga Urinaria/patologíaRESUMEN
Ras is a key regulator of the MAP kinase-signaling cascade and may cause morphologic change of Ras-transformed cells. Signal transducer and activator of transcription 3 (Stat3) can be activated by cytokine stimulation. In this study, we unravel that Ha-ras(V12) overexpression can downregulate the expression of Stat3 protein at a posttranslational level in NIH3T3 cells. Furthermore, we demonstrate that Stat3 expression downregulated by Ha-ras(V12) overexpression is through proteosome degradation and not through a mTOR/p70S6K-related signaling pathway. The suppression of Stat3 accompanied by the morphologic change induced by Ha-ras(V12) was through mitogen extracellular kinase (MEK)/extracellular-regulated kinase (ERK) signaling pathway. Microtubule disruption is involved in Ha-ras(V12)-induced morphologic change, which could be reversed by overexpression of Stat3. Taken together, we are the first to demonstrate that Stat3 protein plays a critical role in Ha-ras(V12)-induced morphologic change. Oncogenic Ras-triggered morphologic change is through the activation of MEK/ERK to posttranslationally downregulate Stat3 expression. Our finding may shed light on developing novel therapeutic strategies against Ras-related tumorigenesis.
Asunto(s)
Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Microtúbulos/ultraestructura , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Proteína Oncogénica p21(ras)/metabolismo , Biosíntesis de Proteínas , Factor de Transcripción STAT3/antagonistas & inhibidores , Animales , Regulación hacia Abajo , Sistema de Señalización de MAP Quinasas , Ratones , Microtúbulos/metabolismo , Células 3T3 NIH , Proteína Oncogénica p21(ras)/genética , Proteínas Quinasas/metabolismo , Proteínas Quinasas S6 Ribosómicas 70-kDa/metabolismo , Serina-Treonina Quinasas TORRESUMEN
The retinoblastoma suppressor (Rb)-associated protein 46 (RbAp46) is a nuclear protein of 46 kDa and contains four repeats that end with Trp-Asp (WD) residues. In this study, we reveal that the RbAp46 protein level upon SUMO-1 expression was increased. The increasing level of RbAp46 protein by SUMO-1 was not regulated at the transcriptional level. SUMO-1 does not affect the degradation of RbAp46. Co-localization of RbAp46 and SUMO-1 in the nuclei of stable NIH/3T3 cells harboring the inducible Ha-ras(Val12) oncogene (pSVlacOras) designated as 7-4, and protein-protein interaction between RbAp46 and SUMO-1 was also detected by co-immunoprecipitation in these cells. However, SUMO-l-related sumoylation was not involved in the modification of RbAp46. It is possibly that SUMO-1 acts through formation of complex with RbAp46 to stabilize RbAp46 protein. Overexpression of RbAp46 protein suppressed the NIH/3T3 cell growth induced by Ha-Ras(V12). SUMO-1 further enhances the suppression of cell growth through stabilization of RbAp46 protein. This is the first report to demonstrate that SUMO-1 can suppress Ras-related cell proliferation through stabilization of RbAp46 protein.