RESUMEN
Paeoniae Radix (PR) is the root of traditional Chinese Herb named Paeonia lactiflora Pallas, which is commonly used to treat liver diseases in China for centuries. Several earlier studies have indicated that PR has anticancer growth activities, however the mechanism underlying these activities was unclear and remained to be elucidated. In this study, we evaluated the molecular mechanism of the effect of PR on human hepatoma cell lines, HepG2 and Hep3B. Our results showed that the water-extract of Paeoniae Radix (PRE) had inhibitory effect on the growth of both HepG2 and Hep3B cell lines. The induction of internucleosomal DNA fragmentation and chromatin condensation appearance, and accumulation of sub-G1 phase of cell cycle profile in PRE treated hepatoma cells evidenced that the cytotoxicity of PRE to the hepatoma cells is through activation of the cell death program, apoptosis. The activation of apoptosis by PRE is independent of the p53 pathway as Hep3B cell is p53-deficient. In addition, the differential gene expression of PRE treated HepG2 was examined by cDNA microarray technology and RT-PCR analysis. We found that the gene expression of BNIP3 was up-regulated while ZK1, RAD23B, and HSPD1 were down-regulated during early apoptosis of the hepatoma cell mediated by PRE. The elucidation of the drug targets of PR on inhibition of tumor cells growth should enable further development of PR for liver cancer therapy.
Asunto(s)
Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Carcinoma Hepatocelular/patología , Medicamentos Herbarios Chinos , Paeonia , Proteínas Proto-Oncogénicas , Factores de Transcripción , Células Tumorales Cultivadas/efectos de los fármacos , Proteína p53 Supresora de Tumor/metabolismo , Proteínas Supresoras de Tumor , Carcinoma Hepatocelular/genética , Chaperonina 60/efectos de los fármacos , Chaperonina 60/genética , Fragmentación del ADN/efectos de los fármacos , Enzimas Reparadoras del ADN , ADN de Neoplasias/análisis , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Humanos , Factores de Transcripción de Tipo Kruppel , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Proteína p53 Supresora de Tumor/deficiencia , Proteína p53 Supresora de Tumor/genéticaRESUMEN
Androgen receptor (AR) is a ligand-dependent transcription factor and its activity is regulated by numerous AR coregulators. Aberrant expression of AR coregulators in prostate cancer cells has an important role in the development and progression of prostate cancer. We report here that CDC25A, a cell cycle-promoting phosphatase over-expressed in a number of cancers, functions as an AR coregulator suppressing the AR transcriptional activity. In this study, we found that CDC25A is upregulated in human prostate cancer and its expression level is positively associated with the Gleason score and disease metastasis. More importantly, we showed that CDC25A can physically interact with AR through its putative catalytic domain. In addition, ectopic expression of CDC25A in prostate cancer cell lines suppresses PSA and Probasin promoter activities significantly, indicating that CDC25A may function as an AR corepressor. This was further confirmed by knockdown of endogenous CDC25A expression using small interfering RNA (siRNA), which resulted in upregulation of PSA promoter activity. Moreover, a truncated mutant that does not interact with AR fails to suppress the PSA promoter activity, indicating that CDC25A downregulates androgen-responsive promoter by physically interacting with AR. Taken together, our results demonstrated a novel function of CDC25A in the regulation of androgen signaling in human prostate cancer cells.
Asunto(s)
Antagonistas de Receptores Androgénicos , Neoplasias de la Próstata/fisiopatología , Proteínas Represoras/metabolismo , Fosfatasas cdc25/metabolismo , Proteína de Unión a Andrógenos/biosíntesis , Línea Celular , Técnicas de Silenciamiento del Gen , Silenciador del Gen , Humanos , Masculino , Antígeno Prostático Específico/biosíntesis , Dominios y Motivos de Interacción de Proteínas , Mapeo de Interacción de Proteínas , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismoRESUMEN
Id-1 is a member of the HLH protein family that regulates a wide range of cellular processes such as cell proliferation, apoptosis, senescence and overexpression of Id-1 was recently suggested to play roles in the development and progression of different cancers. Previously, Id-1 was shown to physically interact with the viral protein E1A. Meanwhile, Id-1 expression was found to be regulated by several of the virus-encoded proteins, suggesting that Id-1 may be a common cellular target of the viral proteins. Here, we report that Id-1 interacts with the Hepatitis-B virus (HBV)-encoded protein HBX and regulates its stability in hepatocellular carcinoma (HCC) cells. We found that in HCC cells, ectopic Id-1 expression significantly decreased the half-life of the HBX protein, indicating that HBX is destabilized by Id-1. Meanwhile, the Id-1-induced HBX degradation was found to be inhibited by treatment with proteasome inhibitor, suggesting that this process is mediated through the proteasome pathway. Interestingly, while Id-1 did not induce HBX-ubiquitination, we found that removal of all the lysine residues of the HBX protein protects it from the effect of Id-1, indicating that ubiquitination is still required for the Id-1-mediated HBX degradation. Meanwhile, we found that Id-1 binds to the proteasome subunit C8 and facilitates its interaction with the HBX protein and disruption of this interaction completely abolishes the negative effect of Id-1 on HBX protein stability. Taken together, our results demonstrated a novel function of Id-1 in regulating HBX protein stability through interaction with the proteasome.