RESUMEN
The promyelocytic leukemia (PML) gene is a tumor suppressor gene. It was first identified in acute promyelocytic leukemia, in which it is fused to retinoic acid receptor α by the (15;17) chromosomal translocation. The function of the PML protein is frequently lost or aberrant in human solid tumors. In human ovarian carcinoma tissue, PML detected by immunohistochemistry was highly expressed. A PML-silencing vector, pSRG-shPml, was constructed and used to transfect human ovarian cancer cells. Cells were cultured and selected with puromycin for 10-15 days, and then the PML mRNA expression levels were detected by RT-qPCR and immunofluorescence. Proliferation and clone number of PML-depleted cells were detected using MTT assay and colony-forming assay. The protein expression associated with DNA damage and apoptosis was assessed in PML-depleted cells using western blot analysis and immunofluorescence. The results showed that PML was highly expressed in human ovarian tissue. The proliferation and colony formation of ovarian cancer cells were significantly inhibited after PML was depleted. Western blot analysis and immunofluorescence revealed that p-H2AX and cleaved caspase-3 expression significantly increased after PML silencing. PML was located in the nucleus, and it formed foci after X-ray irradiation. PML foci increased significantly with increasing irradiation doses.
RESUMEN
Mouse ovarian surface epithelium (OSE) is a single layer of cubodial epithelial cells that covers the ovary surface and is involved in regulating the secretion and transport of 17ß-hydroxysteroid dehydrogenase. Recently, OSE cells have attracted particular interest as a major source of ovarian cancer. Death-associated protein DAXX along with PML (promyelocytic leukemia protein) nuclear bodies (PML-NBs) reportedly play roles in transcriptional regulation and apoptosis. However, little is known regarding a role for DAXX in mOSE cells. In this study, we both over-expressed DAXX and depleted DAXX in primary mOSE cells. We found that Daxx deletion accelerated senescence in a p53/p21-dependent manner and promoted DNA damage by interacting with PML bodies without affecting cell cycle progression. These results suggest that DAXX may transform mOSE cells to an ovarian oncogenic phenotype and may be an anti-cancer target.
Asunto(s)
Proteínas Portadoras/genética , Senescencia Celular/genética , Daño del ADN , Células Epiteliales/metabolismo , Silenciador del Gen , Péptidos y Proteínas de Señalización Intracelular/genética , Proteínas Nucleares/genética , Ovario/metabolismo , Animales , Carcinoma Epitelial de Ovario , Proteínas Portadoras/metabolismo , Ciclo Celular/genética , Proteínas Co-Represoras , Femenino , Eliminación de Gen , Expresión Génica , Humanos , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Ratones , Chaperonas Moleculares , Neoplasias Glandulares y Epiteliales/genética , Neoplasias Glandulares y Epiteliales/metabolismo , Proteínas Nucleares/metabolismo , Proteína Oncogénica p21(ras)/genética , Proteína Oncogénica p21(ras)/metabolismo , Neoplasias Ováricas/genética , Neoplasias Ováricas/metabolismo , Ovario/citología , Proteína de la Leucemia Promielocítica , Factores de Transcripción/metabolismo , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismo , Proteínas Supresoras de Tumor/metabolismoRESUMEN
In Drosophila melanogasler,the wings apart-like (wapl) gene encodes a protein that regulates heterochromatin structure. Human wapl gene is human homolognue of wapl,they have same function. HWAPL is a cohesion-binding protein that affect chromosomal segregation. It has a character of oncoproteins and linked to cervical carcinogenesis and tumor progression.
Asunto(s)
Proteínas Portadoras/genética , Proteínas Portadoras/fisiología , Proteínas de Ciclo Celular/metabolismo , Cromátides/metabolismo , Proteínas Cromosómicas no Histona/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/fisiología , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas/fisiología , Animales , Proteínas Cromosómicas no Histona/genética , Proteínas Cromosómicas no Histona/fisiología , Proteínas de Drosophila/genética , Proteínas de Drosophila/fisiología , Femenino , Humanos , Neoplasias del Cuello Uterino/genética , CohesinasRESUMEN
The recombined adenovirus DNA was transfected into 293 cells for packing and amplification of replication-deficient Ad-CMV-E6/E7, Ad-K14 -E6/E7 virus was purified by CsCl density gradient centrifugation , recombined adenovirus Ad-CMV-E6/E7, Ad-K14 -E6/E7 were used as experimental group, while pAd-CMV and pAdtrack-K14 were used as control group. Four of them were injected through one main vein of nude mice tail respectively. These mice were then treated with 0.05 mg 17beta-estradiol over 12 weeks. Mice were anaesthesiaed with 2.5% Avertint and the vagina, mammary gland, ovaries and uterus were dissected and fixed in 3.75% paraformaldehyde overnight at 4 degrees C. Paraffin-embedded sections, HE staining and identification of P53 and Bcl-2 protein via immunohistochemistry were performed. The expression of E6/E7 was verified by RT-PCR in different tissue of nude mice. HE staining showed evident hyperplasy in cervix-uterus transformation zone of experimental group 2. The expression of mutant P53 and Bcl-2 were higher than control group via immunohistochemical S-P method in uterus stroma-cell. Western blotting also showed that E6 protein was expressed. The expression of E6/E7 was higher than control group by human cytokeratin promoter 14 and hyperlasy changes were detected in epithelial tissue of cervix-uterus transformation zone.