The H19 TATA-less promoter is efficiently repressed by wild-type tumor suppressor gene product p53
The H19 TATA-less promoter is efficiently repressed by wild-type tumor suppressor gene product p53
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DOI：
10.1038/sj.onc.1201742
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发表时间：
1998-05-07
期刊：
影响因子：
8
通讯作者：
Curgy, JJ
中科院分区：
文献类型：
--
作者：
Dugimont, T;Montpellier, C;Curgy, JJ
发育调节的H19基因显示出几个显著的特性：明显非翻译的mRNA的表达，基因组印记（仅表达母体等位基因），印记的松弛和/或在一些肿瘤中表现出的表观遗传病变。尽管放松后的印记状态的基因的几个观察，数据的反式和顺式作用的因素所需的人H19基因的表达仍然是失踪。作为第一种方法来解决参与基因调控的因素的识别，我们发现，从p53反义转染的HeLa克隆细胞显示H19转录物的量增加时，与未转染的细胞相比。此外，HeLa克隆稳定转染温度敏感（TS）143 Ala p53突变体表现出温度依赖性调节H19的表达。p53蛋白对H19表达的抑制作用的这种初步指示已经通过在HeLa细胞中的瞬时共转染实验得到证实，所述实验使用在H19基因的转录起始点的紧邻上游的823 bp序列的控制下的荧光素酶替代构建体，以及含有正义、反义或ts 143 Ala突变体p53 cDNA的不同构建体。我们观察到在非允许温度下，反义p53 cDNA和温度敏感突变体p53在瞬时共转染中H19启动子驱动的活性增加，但正义野生型p53 cDNA的活性降低。此外，在缺乏内源性p53的细胞系（Calu 6细胞）中重复共转染实验，结果为p53蛋白下调H19基因的表达提供了额外的证据。
The developmentally regulated H19 gene displays several remarkable properties: expression of an apparently non-translated mRNA, genomic imprinting (maternal allele only expressed), relaxation of the imprinting and/or epigenetic lesions demonstrated in some tumors. Despite several observations after relaxation of imprinting status of the gene, data on tr ans and cis-acting factors required for the human H19 gene expression are still missing. As a first approach to address identification of factors involved in the regulation of the gene, we found that cells from a p53 antisense-transfected HeLa clone displayed increased amounts of H19 transcripts when compared to the non-transfected cells. Moreover, a HeLa clone stably transfected with a temperature sensitive (ts) 143 Ala p53 mutant exhibited temperature-dependent regulation of H19 expression. This preliminary indication of the repressing effect of the p53 protein on H19 expression has been confirmed by transient cotransfection experiments in HeLa cells, using luciferase surrogate constructs under the control of the 823 bp sequence immediately upstream of the transcription start point of the H19 gene, and different constructs containing sense, antisense or a ts 143 Ala mutant p53 cDNA. We observed an increase of H19 promoter-driven activity in transient cotransfections with the antisense p53 cDNA and the temperature sensitive mutant p53 at the non-permissive temperature, but a decrease with sense wild-type p53 cDNA. Furthermore, the cotransfection experiments were repeated in a cell line lacking endogenous p53, (Calu 6 cells) and the results provided additional evidence for a down regulation of the expression of the H19 gene by the p53 protein.