TRANSCRIPTIONAL REGULATION OF CYTOCHROME P-450 GENES

许多P450基因在啮齿动物肝脏中结构性表达。有些人， 例如大鼠的CYP2EI和CYP2AI基因，直到出生时才会消失 当它们被转录激活的时候。CYP2EL仍然活跃在 男性和女性在整个成年生活中都是如此，而CYP2A1基因 在男性青春期受到抑制，但在女性体内保持活跃。A相关 基因，CYP2A2，在女性的一生中转录不活跃 但在青春期男性会变得活跃，同时会抑制 细胞色素P4 2 AI基因。大鼠的CYP2D基因也在某些阶段被激活 发展。CYP2D3在出生后立即被激活，而CYP2D5在出生后立即被激活 男女都在青春期被激活。探讨其发病机制。 基因的调控，我们正在使用的程序可以识别 顺式作用的DNA元件和交易因素 转录激活。在体外，细胞自由转录系统是 从不同年龄大鼠的肝脏制备并用于转录克隆 P450基因模板。进行删除分析以确定重要的 RNA聚合酶II启动子上游的调控区。DNA结合 分析用于鉴定能够结合的顺式作用元件 核转录因子。使用这些程序，我们已经确定 控制细胞色素P450酶EI发育激活的因素 吉恩。细胞色素P450 2 A基因的调控机制仍不清楚。 对P450的诱导机制的研究也在进行中 外来化合物的基因。克隆了细胞色素P4AI和细胞色素P4A2基因 完全测序。这些基因可以被转录激活 降血脂药氯贝特。CYP4A基因的启动子区域为 与报告基因氯霉素乙酰转移酶融合。转染腺病毒载体 这些构建物进入大鼠肝癌细胞或大鼠肝细胞中不产生 氯贝特治疗后的活动度。进一步的研究正在进行中 使用基因上游高达20千碱基对的DNA片段 寻找能够激活异源启动子的序列的起始站点 在氯贝特存在的情况下。

A number of P450 genes are constitutively expressed in rodent liver. Some, such as the rat CYP2EI and CYP2AI genes, are absent until birth at which time they become transcriptionally activated. The CYP2El remains active in both male and female throughout adult life, whereas the CYP2Al gene is repressed in males at puberty but remains active in females. A related gene, CYP2A2, is transcriptionally inactive throughout the life of females but becomes active in males at puberty concomitant with suppression of the CYP2AI gene. The rat CYP2D genes are also activated at certain stages of development. CYP2D3 is activated right after birth, while CYP2D5 is activated at puberty in both sexes. To investigate the mechanism of regulation of the genes, we are using procedures that can identify cis-acting DNA elements and transacting factors responsible for transcriptional activation. In vitro cell free transcription systems are prepared from livers of rats of various ages and used to transcribe cloned P450 gene template. Deletion analyses are undertaken to identify important regulatory regions upstream of the RNA polymerase II promoter. DNA binding analyses are used to identify cis-acting elements capable of binding nuclear transcription factors. Using these procedures we have identified the elements and factors controlling developmental activation of the CYP2EI gene. The mechanism of regulation of the CYP2A genes remains elusive. Studies are also underway to determine the mechanism of induction of P450 genes by foreign compounds. The CYP4AI and CYP4A2 genes were cloned and completely sequenced. These genes can be transcriptionally activated by the hypolipidemic drug clofibrate. The promoter regions of CYP4A genes were fused to the report gene chloramphenicol acetyltransferase. Transfection of these constructs into rat hepatoma cells or rat hepatocytes yielded no activity after treatment with clofibrate. Further studies are being carried out using fragments of DNA up to 20 kilobase pairs upstream of the gene's start site to search for sequences that can activate heterologous promoters in the presence of clofibrate.