摘要： Hodgkin/Reed-Sternberg (HRS) cells of classical Hodgkin lymphoma (cHL) display unique characteristics that discriminate cHL from other B-cell lymphomas and normal B cells. Therefore, comparative gene expression profiling of Hodgkin and non-Hodgkin B cells could lead to the identification of candidate genes that are critical for the pathogenesis of cHL. We performed microarray analysis of Hodgkin and non-Hodgkin cell lines and identified activating transcription factor 3 (ATF3), a member of the cyclic AMP response element binding protein (CREB)/ATF family, as a differentially expressed candidate gene. Extensive analysis of a large panel of cell lines, primary tumor samples, and normal tissues revealed that high expression of ATF3 is found in nearly all cases of cHL and is almost exclusively restricted to it. Selective knock-down of ATF3 by RNA interference suppressed proliferation and strongly reduced viability of Hodgkin cells. Thus, overexpression of ATF3 is a molecular hallmark of cHL that contributes to the malignant growth of HRS cells.

关键词： ATF3   activating transcription factor 3   BO   Barrett's oesophagus   BEBM   bronchial epithelial basal media   DAF   decay accelerating factor   EGRF (early growth response factor   EGR1   early growth response gene 1   GORD   gastro-oesophageal reflux disease   IL-8   interleukin-8   NF-KB   nuclear factor-kappa-B   NR4A2   nuclear receptor 4A2   PTGS2   prostaglandin synthase-2   TFBS   transcription factor binding site  

摘要： The development of gastro-oesophageal reflux disease (GORD) is known to be a causative risk factor in the evolution of adenocarcinoma of the oesophagus. The major component of this reflux is gastric acid. However, the impact of low pH on gene expression has not been extensively studied in oesophageal cells. This study utilizes a transcriptomic and bioinformatic approach to assess regulation of gene expression in response to low pH. In more detail, oesophageal adenocarcinoma cell lines were exposed to a range of pH environments. Affymetrix microarrays were used for gene-expression analysis and results were validated using cycle limitation and real-time RT-PCR analysis, as well as northern and western blotting. Comparative promoter transcription factor binding site (TFBS) analysis (MatInspector) of hierarchically clustered gene-expression data was employed to identify the elements which may co-ordinately regulate individual gene clusters. Initial experiments demonstrated maximal induction of EGR1 gene expression at pH 6.5. Subsequent array experimentation revealed significant induction of gene expression from such functional categories as DNA damage response (EGR1-4, ATF3) and cell-cycle control (GADD34, GADD45, p57). Changes in expression of EGR1, EGR3, ATF3, MKP-1, FOSB, CTGF and CYR61 were verified in separate experiments and in a variety of oesophageal cell lines. TFBS analysis of promoters identified transcription factors that may co-ordinately regulate gene-expression clusters, Cluster 1: Oct-1, AP4R;Cluster 2: NF-kB, EGRF;Cluster 3: IKRS, AP-1F. Low pH has the ability to induce genes and pathways which can provide an environment suitable for the progression of malignancy. Further functional analysis of the genes and clusters identified in this low pH study is likely to lead to new insights into the pathogenesis and therapeutics of GORD and oesophageal cancer.

摘要： Background: This is a "proof of concept study" to test the hypothesis that pulsed radiofrequency, PRF, produces cell stress at the primary afferent level without signs of overt thermal damage. We assumed that cell stress would result in impairment of normal function, and used the expression of activating transcription factor 3, ATF3, as an indicator of cellular "stress". Methods: PRF (20 ms of 500-kHz RF Pulses, delivered at a rate of 2 Hz;maximum temperature 42 degrees C) was delivered either to the sciatic nerve of adult rats in mid thigh, or to the L4 anterior primary ramus just distal to the intervertebral foramen. Controls were sham-operated or L4 axotomised. All tissues were examined 14 days after Surgery. The percentage of CGRP- or ATF3-positive DRG neuronal somata was calculated using image analysis software (SigmaScan Pro 4). Results: ATF3 expression was upregulated in L4 DRG neuronal cell bodies, irrespective of their size, after axotomy. It was also upregulated significantly (p<0.002) and selectively, in small and medium calibre L4 DRG neurons, when PRF was applied close to the DRG just distal to the intervertebral foramen. PRF did not produce any obvious cellular changes in the nerve or L4 DRG neurons when applied to the sciatic nerve in mid-thigh. Conclusion: PRF has a biological effect, unlikely to be related to overt thermal damage. It appears to be selective in that it targets the group of neurons whose axons are the small diameter C and A delta nociceptive fibres. (C) 2005 European Federation of Chapters of the International Association for the Study of Pain. Published by Elsevier Ltd. All rights reserved.

摘要： Previous results of our cDNA microarray analysis to look for genes whose expression level correlates well with in vitro tubulogenesis by NP31 endothelial cells revealed the transcription factor ATF3 known to be responsive to stress such as reactive oxygen species (ROS). Anti-ATF3 small interfering RNA gave an inhibitory influence on tube formation by NP31 cells expressing an activated form of the vascular endothelial growth factor receptor 1 (VEGFR-1) kinase. When expression of ATF3 was regulated under the control of tetracycline system in NP31 cells, they acquired the tubulogenic ability upon ATF3 induction. While ATF3 failed to induce expressions of VEGF and VEGFR, it regulated those of CDK2, CDK4, p8, plasminogen activator inhibitor 1, integrin alpha 1, subunit and matrix metalloprotease MMP13. In H2O2-stimulated NP31 cells as well as endothelial cells of glomerulus and aorta of Otsuka-Long-Evans-Tokushima-Fatty diabetic model rats, concomitantly enhanced expressions of ATF3, PAI-1, and p8 were observed. Given the proposed hypothesis of the close linkage between diabetic angiopathy and ROS, those data suggest that ROS-associated diabetic complication may involve ATF3-mediated pathological angiogenesis.

关键词： Genes  

摘要： Activating transcription factor (ATF) 3 plays a role in determining cell fate and generates a variety of alternatively spliced isoforms in stress response. We have reported previously that splice variant ATF3 Delta Zip2, which lacks the leucine zipper region, is induced in response to various stress stimuli. However, its biological function has not been elucidated. By using cells treated with tumor necrosis factor-alpha and actinomycin D or cells overexpressing ATF3 Delta Zip2, we showed that ATF3 Delta Zip2 sensitizes cells to apoptotic cell death in response to tumor necrosis factor-alpha, at least in part through suppressing nuclear factor (NF)-kappa B-dependent transcription of antiapoptotic genes such as cIAP2 and XIAP. ATF3 Delta Zip2 interacts with a p65 (RelA)-cofactor complex containing CBP/p300 and HDAC1 at NF-kappa B sites of the proximal promoter region of the cIAP2 gene in vivo and down-regulates the recruitment of CBP/p300. Our study revealed that ATF3 Delta Zip2 counteracts anti-apoptotic activity of NF-kappa B, at least in part, by displacing positive cofactor CBP/p300 and provides insight into the mechanism by which ATF3 regulates cell fate through alternative splicing in stress response.

关键词： axotomy   neuropathic pain   rat   sham   transcription factor  

摘要： Activating transcription factor 3 (ATF3) is a widely used marker of damaged primary sensory neurons that is induced in essentially all dorsal root ganglion (DRG) neurons by spinal nerve axotomy. Whether such injuries induce its expression in neurons of adjacent DRGs remains unknown. Following L5 spinal nerve ligation, experimental but not sham-operated rats develop thermal and mechanical hypersensitivity. In the L4 DRG, 11-12% of neurons were ATF3 positive by 1 day post-surgery, and numbers remain unchanged at 2 weeks. Importantly, sham exposure of the L5 spinal nerve produced a nearly identical number of ATF3-positive neurons in the L4 DRG and also a substantial increase in the L5 DRG, with a similar time-course to experimental animals. There was no correlation between behaviour and magnitude of ATF3 expression. Co-localization studies with the DRG injury markers galanin, neuropeptide Y and nitric oxide synthase (NOS) showed that approximately 75, 50 and 25%, respectively, of L4 ATF3-positive neurons co-expressed these markers after L5 transection or sham surgery. Additionally, increases in galanin and NOS were seen in ATF3-negative neurons in L4. Our results strongly suggest that the surgical exposure of spinal nerves induces ATF3 in the L4-5 DRG, irrespective of whether the L5 nerve is subsequently cut. This probably reflects minor damage to the neurons or their axons but nevertheless is sufficient to induce phenotypic plasticity. Caution is therefore warranted when interpreting the phenotypic plasticity of DRG neurons in adjacent ganglia in the absence of positive evidence that they are not damaged.

关键词： Restin   ATF3   Cyclin D1   报告系统   荧光素酶   α-32P标记的探针   点杂交  

摘要： Restin是1999年本校生物化学与分子生物学教研室在研究全反式维甲酸诱导细胞分化时克隆得到的一个未知功能的人类新基因，2001年，Chomez等通过同源序列筛选Genebank数据库，也发现了该基因，并命名为MAGE-H1，该基因主要表达于终末分化细胞。稳定转染该基因的细胞出现G1期阻滞，抑制细胞的增殖，提示Restin与细细胞周期的调控有关。但有关该基因的具体生物学功能研究还未见报道。 实验前期研究利用细胞双杂交技术，对Restin在细胞周期中可能的作用的转录因子(ATF3、ATF4、E2F6、C-JUN、C-FOS、p53)进行了双杂交筛选，结果发现ATF3与Restin表现出很强的相互作用。为了进一步验证ATF3与Restin的相互作用，对ATF3进行截短并构建了一系列ATF3的截短片段的双杂交质粒，再次运用细胞双杂交进行相互作用的探索。结果发现Restin与ATF3的确存在相互作用，并作用于ATF3的DNA结合结构域(ATF3 binding domain，ATF3／BD)，不完整的ATF3／BD降低了Restin与ATF3相互作用能

关键词： 单纯性尿道下裂   SRD5A2   ATF3   RT-PCR   免疫组化   基因突变   PCR-SSCP  

摘要： 尿道下裂主要表现为异位的尿道外口，在男性新生儿中其发病率约为1／250～300，它是男性泌尿生殖系统最常见先天性畸形之一。近30年来尿道下裂的发病率呈逐年升高趋势，大部分是散发的、无遗传和家族史。既往研究显示尿道下裂存在家族性单基因遗传现象，或伴发于其它的遗传疾病综合症，同时也有研究发现环境因素与尿道下裂有关，目前认为尿道下裂是遗传和环境因素综合作用的结果。 胎儿尿道发育有赖于足量的雄性激素，它包括睾酮和双氢睾酮，睾酮在由SRD5A2基因编码的Ⅱ型5α-还原酶的作用下转化成活性更高的双氢睾酮。研究证明Ⅱ型5α-还原酶的功能异常、SRD5A2基因突变和尿道下裂的发生有密切关系，至今已发现约40种突变类型，大部分表现为误义突变和无义突变。以往对该基因的研究主要集中在家族性和伴发于其它遗传疾病综合症的尿道下裂，对散发的单纯性尿道下裂研究较少。 ATF3是碱性亮氨酸拉链区（bZIP）家族中ATF／CREB转录因子亚家族的成员之一。研究发现ATF3通过复杂的机制调节其靶基因的表达，它与内环境稳定、伤口愈合、细胞粘附、肿瘤细胞浸润和信号转导通路有关。2003年，Wang等采用微阵列技术发现ATF3是尿道下裂相关基因之一，提示ATF3基因除了对应激产生保护性反应外，可能与尿道下裂发生过程中的应激应答有关。进一步研究ATF3和散发的单纯性尿道下裂的关系，有可能揭示ATF3在其发病中的作用机制。 家族性尿道下裂约占7％，其余的大部分为散发的单纯性尿道下裂。