Biochemical analysis of the human vs. mouse Ah receptor

描述(由申请人提供)：在啮齿动物模型中，AH受体(AhR)已被证明与2，3，7，8-四氯对二恶英(TCDD)的致癌和毒性特性有关。人类人群暴露于低剂量的TCDD，以及其他一些AhR配体，暴露的实际健康影响仍有待确定。老鼠模型中产生的数据能否有效地外推到人类身上，将需要更好地理解物种之间潜在的遗传差异。需要检验的中心假设是，人的AhR在生化上不同于小鼠的AhR，这些差异导致AhR介导的活性改变。拟议研究的另一个目标是更好地了解AhR的多种调节机制。中心假说得到了我们实验室的初步发现的支持。在这一应用中，将对人AhR的生化和转录活性进行表征，并与小鼠AhR(S)进行比较。我们提出了三个特定的目标：L)确定核心未连接受体复合体中人和鼠AhR受体的生化行为，并检测XAP2改变人和鼠AhR活性的能力；2)通过使用嵌合受体来检测人和鼠AhR的反式激活潜力、共激活子募集的特异性和核质穿梭特性；以及3)比较连接的人和小鼠AhR在AhR缺失的tsSV-40转化的小鼠肝细胞系中改变基因表达的能力。在这些目标中，将在同一细胞系中研究小鼠AhR和人类AhR，以允许直接确定物种差异，而不受先前研究中存在的细胞环境变化的影响。总而言之，这些研究将确定人类和小鼠AhR功能的差异水平，并应该能够更好地评估人类接触TCDD的意义。

DESCRIPTION (provided by applicant): The Ah receptor (AhR) has been shown to be responsible for the carcinogenic and toxic properties of 2,3,7,8- tetrachloro-p-dioxin (TCDD) in rodent models. The human population is exposed to low doses of TCDD, and a number of other AhR ligands, the actual health effects of exposure remain to be established. Whether or not data generated in the mouse model can be effectively extrapolated to humans will require a better understanding of potential genetic differences between species. The central hypothesis to be tested is that the human AhR is biochemically different from the mouse AhR and these differences result in altered AhR-mediated activity. An additional goal of the proposed studies is to better understand the multiple mechanisms of regulation of the AhR. The central hypothesis is supported by preliminary findings from our laboratory. In this application the biochemical and transcriptional activity of the human AhR will be characterized and compared with the mouse AhR(s). Three specific aims are proposed; l) Determine the biochemical behavior of the human vs. mouse Ah receptor in the core unliganded receptor complex, and examine the ability of XAP2 to alter the activity of the human vs. mouse AhR, 2) Examine the transactivation potential, co-activator recruitment specificity and nucleocytoplasmic shuttling properties of the human AhR compared with the mouse AhR through the use of chimeric receptors, and 3) Compare the ability of the liganded human AhR vs. the mouse AhR to alter gene expression in AhR-null tsSV-40 transformed mouse hepatocyte cell lines. In these aims the mouse AhR and human AhR will be studied in the same cell line to allow a direct determination of species differences independent of cell context variation present in previous studies. Collectively, these studies will establish the level of divergence in human vs. mouse AhR function and should allow a better assessment of the significance of human exposure to TCDD.