CATHEPSIN-D GENE IS CONTROLLED BY A MIXED PROMOTER, AND ESTROGENS STIMULATE ONLY TATA-DEPENDENT TRANSCRIPTION IN BREAST-CANCER CELLS
CATHEPSIN-D GENE IS CONTROLLED BY A MIXED PROMOTER, AND ESTROGENS STIMULATE ONLY TATA-DEPENDENT TRANSCRIPTION IN BREAST-CANCER CELLS
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DOI：
10.1073/pnas.90.1.203
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发表时间：
1993-01-01
影响因子：
11.1
通讯作者：
ROCHEFORT, H
中科院分区：
文献类型：
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作者：
CAVAILLES, V;AUGEREAU, P;ROCHEFORT, H
组织蛋白酶1（cath-D）基因编码一种普遍存在的溶酶体乙酰化蛋白酶，在侵袭性人类乳腺癌中过表达，并且其转录在对雌激素敏感的乳腺癌细胞中由雌激素诱导。我们已经确定了来自MCF 7细胞的人cath-D基因的近5'上游区的结构和功能。我们发现，该启动子具有一个复合结构，具有管家基因（高G + C含量和潜在的转录因子Spl位点）和调控基因（TATAA序列）的特征。通过RNA酶保护实验，我们发现转录起始于5个主要转录位点（TSSI至-V），跨越52个碱基对。在对雌二醇有反应的乳腺癌细胞中，雌二醇使TSSI起始的RNA水平增加了6- 10倍，TSSI位于TATA盒下游约28个碱基对处。特异性调节雌二醇的转录开始在网站I完全证实了引物延伸。此外，在ZR 75 -1细胞系和稳定转染雌激素受体的MDA-MB 231雌激素抗性乳腺癌细胞中观察到相同的雌二醇效应。定点突变表明，TATA盒是必不可少的启动cath-D基因转录在TSSI。在乳腺癌活检样本中，高水平的TATA依赖性转录与cath-D mRNA的过表达相关。我们的结论是，cath-D的行为，根据条件，作为一个管家基因与多个起始位点或作为一个可控制的TATA盒的酶调节基因。
The cathepsin 1) (cath-D) gene, coding for a ubiquitous lysosomal aspartyl protease, is overexpressed in aggressive human breast cancers, and its transcription is induced by estrogens in hormone-responsive breast cancer cells. We have determined the structure and function of the proximal 5' upstream region of the human cath-D gene from MCF7 cells. We show that the promoter has a compound structure with features of both housekeeping genes (high G + C content and potential transcription factor Spl sites) and regulated genes (TATAA sequence). By RNase protection assay, we show that transcription is initiated at rive major transcription sites (TSSI to -V) spanning 52 base pairs. In hormone-responsive breast cancer cells, estradiol increased by 6- to 10-fold the level of RNAs initiated at TSSI, which is located about 28 base pairs downstream from the TATA box. The specific regulation by estradiol of transcription starting at site I exclusively was confirmed by primer extension. Moreover, the same estradiol effect was observed in the ZR75-1 cell line and in MDA-MB231 estrogen-resistant breast cancer cells stably transfected with the estrogen receptor. Site-directed mutagenesis indicated that the TATA box is essential for initiation of cath-D gene transcription at TSSI. In breast cancer biopsy samples, high levels of TATA-dependent transcription were correlated with overexpression of cath-D mRNA. We conclude that cath-D behaves, depending on the conditions, as a housekeeping gene with multiple start sites or as a hormone-regulated gene that can be controlled from its TATA box.