Varanasi U, Chu R, Huang Q, Castellon R, Yeldandi A V, Reddy J K
Department of Pathology, Northwestern University Medical School, Chicago, Illinois 60611, USA.
J Biol Chem. 1996 Jan 26;271(4):2147-55. doi: 10.1074/jbc.271.4.2147.
Peroxisome proliferators cause a rapid and coordinated transcriptional activation of genes encoding the enzymes of the peroxisomal beta-oxidation pathway in rats and mice. Cis-acting peroxisome proliferator responsive elements (PPREs) have been identified in the 5'-flanking region of H202-producing rat acyl-CoA oxidase (ACOX) gene and in other genes inducible by peroxisome proliferators. To gain more insight into the purported nonresponsiveness of human liver cells to peroxisome volume density and in the activity of the beta-oxidation enzyme system, we have previously cloned the human ACOX gene, the first and rate-limiting enzyme of the peroxisomal beta-oxidation system. We now present information on a regulatory element for the peroxidase proliferator-activated receptor (PPAR)/retinoid X receptor (RXR) heterodimers. The PPRE, consists of AGGTCA C TGGTCA, which is a direct repeat of hexamer half-sites interspaced by a single nucleotide (DR1 motif). It is located at -1918 to -1906 base pairs upstream of the transcription initiation site of this human ACOX gene. This PPRE specifically binds to baculovirus-expressed recombinant rat PPAR alpha/RXR alpha heterodimers. In transient transfection experiments, the maximum induction of luciferase expression by ciprofibrate and/or 9-cis-retinoic acid is dependent upon cotransfection of expression plasmids for PPAR alpha and RXR alpha. The functionally of this human ACOX promoter was further demonstrated by linking it to a beta-galactosidase reporter gene or to a rat urate oxidase cDNA and establishing stably transfected African green monkey kidney (CV1) cell lines expressing reporter protein. The human ACOX promoter has been found to be responsive to peroxisome proliferators in CV1 cells stably expressing PPAR alpha, whereas only a basal level of promoter activity is detected in stably transfected cells lacking PPAR alpha. The presence of a PPRE in the promoter of this human peroxisomal ACOX gene and its responsiveness to peroxisome proliferators suggests that factors other than the PPRE in the 5'-flanking sequence of the human ACOX gene may account for differences, if any, in the pleiotropic responses of humans to peroxisome proliferators.
过氧化物酶体增殖剂可使大鼠和小鼠体内编码过氧化物酶体β-氧化途径中酶的基因迅速且协调地转录激活。在产生H2O2的大鼠酰基辅酶A氧化酶（ACOX）基因的5'侧翼区域以及其他可被过氧化物酶体增殖剂诱导的基因中，已鉴定出顺式作用的过氧化物酶体增殖剂反应元件（PPREs）。为了更深入了解人类肝细胞对过氧化物酶体体积密度以及β-氧化酶系统活性的所谓无反应性，我们之前克隆了人类ACOX基因，它是过氧化物酶体β-氧化系统的首个且限速酶。我们现在展示关于过氧化物酶体增殖剂激活受体（PPAR）/视黄酸X受体（RXR）异二聚体的调控元件的信息。该PPRE由AGGTCA C TGGTCA组成，它是由单个核苷酸间隔的六聚体半位点的直接重复序列（DR1基序）。它位于该人类ACOX基因转录起始位点上游-1918至-1906碱基对处。此PPRE特异性结合杆状病毒表达的重组大鼠PPARα/RXRα异二聚体。在瞬时转染实验中，环丙贝特和/或9-顺式视黄酸对荧光素酶表达的最大诱导依赖于PPARα和RXRα表达质粒的共转染。通过将其与β-半乳糖苷酶报告基因或大鼠尿酸氧化酶cDNA连接，并建立稳定转染的表达报告蛋白的非洲绿猴肾（CV1）细胞系，进一步证明了该人类ACOX启动子的功能。已发现人类ACOX启动子在稳定表达PPARα的CV1细胞中对过氧化物酶体增殖剂有反应，而在缺乏PPARα的稳定转染细胞中仅检测到基础水平的启动子活性。该人类过氧化物酶体ACOX基因启动子中存在PPRE及其对过氧化物酶体增殖剂的反应性表明，人类ACOX基因5'侧翼序列中除PPRE之外的其他因素可能解释了人类对过氧化物酶体增殖剂多效性反应中的差异（如果存在差异的话）。
