雄激素非依赖性前列腺癌细胞雄激素受体靶基因的筛选与初步鉴定

采用染色质免疫共沉淀技术在全基因组水平筛选雄激素非依赖性前列腺癌细胞LNCaP-AI的雄激素受体结合位点,行高通量测序及生物信息学分析共得到2 876个peak（pvalue〈1×10–5）,peak平均长度为673 bp;将peak序列定位到Hg19基因组,共有1 865个靶基因,其中fold enrichment≥10的基因有425个。对peak相关基因进行GO分析发现,与细胞、细胞组分、细胞过程、结合、细胞器相关的基因位列前五位;对peak相关基因进行通路分析发现,与黏着斑、代谢通路、癌症中的转录错误调控、嘌呤代谢等信号通路相关的基因占大多数。筛选出7个候选AR靶基因,采用Real-time qPCR技术分析它们在LNCaP-AI细胞和雄激素依赖性前列腺癌细胞LNCaP中对DTH刺激的反应性,发现DHT刺激可改变7个候选AR靶基因在LNCaP-AI细胞中的表达,为进一步研究雄激素依赖性前列腺癌向非依赖性前列腺癌发展的过程中雄激素受体及其调控的下游靶基因功能起着至关重要的作用。     

Neurogenin 3 recruits CBP co-activator to facilitate histone H3/H4 acetylation in the target gene INSM1

INSM1 is a downstream target gene of neurogenin 3 (ngn3). A promoter construct containing the -426/+40bp region transiently co-transfected into NIH-3T3 cells with a ngn3 expression plasmid resulted in a 12-fold increase in promoter activity. The ngn3/E47 heterodimer selectively binds and activates the E-box3 of the INSM1 promoter. The endogenous ngn3 and CREB-binding protein (CBP) co-activator occupy the INSM1 promoter, resulting in hyper-acetylation of histone H3/H4 chromatin in a human neuroblastoma cell line, IMR-32. Additionally, adenoviral ngn3 can induce endogenous INSM-1 expression in pancreatic ductal carcinoma-1 cells through the recruitment of CBP to the INSM1 promoter and increase the acetylation of the INSM1 promoter region.     

ATP hydrolysis is essential for Bag-1M-mediated inhibition of the DNA binding by the glucocorticoid receptor

The 70-kDa heat shock protein (Hsp70) is involved in providing the appropriate conformation of various nuclear hormone receptors, including the glucocorticoid receptor (GR). The Bcl-2 associated athanogene 1M (Bag-1M) is known to downregulate the DNA binding by the GR. Also, Bag-1M interacts with the ATPase domain of Hsp70 to modulate the release of the substrate from Hsp70. In this study, we demonstrate that ATP hydrolysis enhances Bag-1M-mediated inhibition of the DNA binding by the GR. However, the inhibitory effect of Bag-1M was abolished when the intracellular ATP was depleted. In addition, a Bag-1M mutant lacking the interaction with Hsp70 did not influence the GR to bind DNA, suggesting the interaction of Bag-1M with Hsp70 in needed for its negative effect. These results indicate that ATP hydrolysis is essential for Bag-1M-mediated inhibition of the DNA binding by the GR and Hsp70 is a mediator for this process.     

Induction of human aquaporin-1 gene by retinoic acid in human erythroleukemia HEL cells

Retinoids have been implicated in the control of cell proliferation, differentiation, and developmental processes. We report here that aquaporin-1 (AQP1) is specifically induced by retinoic acid (RA) in human erythroleukemia HEL cells. Both all-trans-RA (ATRA) and 9-cis-RA (9CRA) strongly induced the AQP1 mRNA and protein in a dose-dependent manner. AQP1 protein was mainly expressed in plasma membrane in cells induced by RAs. To identify the RA response element (RARE) in the human AQP1 promoter, the 5(')-flanking region of AQP1 promoter was isolated and transient transfection experiment in HEL cells was performed. Deletion analysis of the AQP1 promoter revealed that one putative DR5-like RARE with five spaces was located in the region from -2218 to -2202; AGGGCAgggacAGGTGA. Electrophoretic mobility shift assay (EMSA) experiment demonstrated that two slowly migrated complexes (C1 and C2) capable of binding the RARE sequence were present in nuclear extracts prepared from cells and the complex C1 was strongly increased in nuclear extracts by RA stimulation. The complexes C1 and C2 were significantly abolished by an excess unlabeled probe. These results indicate that RAs strongly stimulate the human AQP1 gene expression through the RARE and define a novel role in the regulation of erythropoiesis.