Mol P C, Wang R H, Batey D W, Lee L A, Dang C V, Berger S L
Section on Genes and Gene Products, National Cancer Institute, Bethesda, Maryland 20892, USA.
Mol Cell Biol. 1995 Dec;15(12):6999-7009. doi: 10.1128/MCB.15.12.6999.
The Myc protein has been reported to activate transcription of the rat prothymosin alpha gene by binding to an enhancer element or E box (CACGTG) located in the first intron (S. Gaubatz et al., Mol. Cell. Biol. 14:3853-3862, 1994). The human prothymosin alpha gene contains two such motifs: in the promoter region at kb -1.2 and in intron 1, approximately 2 kb downstream of the transcriptional start site in a region which otherwise bears little homology to the rat gene. Using chloramphenicol acetyltransferase (CAT) reporter constructs driven either by the 5-kb human prothymosin alpha promoter or by a series of truncated promoters, we showed that removal of the E-box sequence had no effect on transient expression of CAT activity in mouse L cells. When intron 1 of the prothymosin alpha gene was inserted into the most extensive promoter construct downstream of the CAT coding region, a diminution in transcription, which remained virtually unchanged upon disruption of the E boxes, was observed. CAT constructs driven by the native prothymosin alpha promoter or the native promoter and intron were indifferent to Myc; equivalent CAT activity was observed in the presence of ectopic normal or mutant Myc genes. Similarly, expression of a transiently transfected wild-type prothymosin alpha gene as the reporter was not affected by a repertoire of myc-derived genes, including myc itself and dominant or recessive negative myc mutants. In COS-1 cells, equivalent amounts of the protein were produced from transfected prothymosin alpha genes regardless of whether genomic E boxes were disrupted, intron 1 was removed, or a repertoire of myc-derived genes was included in the transfection cocktail. More importantly, cotransfection of a dominant negative Max gene failed to reduce transcription of the endogenous prothymosin alpha gene in COS cells or the wild-type transfected gene in COS or L cells. Taken together, the data do not support the idea that Myc activates transcription of the intact human prothymosin alpha gene or reporter constructs that mimic its structure. Rather, they suggest that the human prothymosin alpha promoter and downstream elements are buffered so as to respond poorly, if at all, to transient fluctuations in transcription factors which regulate other genes.
据报道，Myc蛋白通过与位于大鼠前胸腺素α基因第一内含子中的增强子元件或E盒（CACGTG）结合来激活该基因的转录（S. Gaubatz等人，《分子与细胞生物学》14:3853 - 3862，1994年）。人类前胸腺素α基因包含两个这样的基序：一个在启动子区域的kb -1.2处，另一个在转录起始位点下游约2 kb的内含子1中，该区域与大鼠基因几乎没有同源性。使用由5 kb人类前胸腺素α启动子或一系列截短启动子驱动的氯霉素乙酰转移酶（CAT）报告构建体，我们发现去除E盒序列对小鼠L细胞中CAT活性的瞬时表达没有影响。当前胸腺素α基因的内含子1插入到CAT编码区下游最完整的启动子构建体中时，观察到转录减少，而在E盒被破坏后转录几乎没有变化。由天然前胸腺素α启动子或天然启动子和内含子驱动的CAT构建体对Myc不敏感；在存在异位正常或突变Myc基因的情况下观察到等效的CAT活性。同样，作为报告基因的瞬时转染野生型前胸腺素α基因的表达不受一系列myc衍生基因的影响，包括myc本身以及显性或隐性负性myc突变体。在COS - 1细胞中，无论基因组E盒是否被破坏、内含子1是否被去除，或者转染混合物中是否包含一系列myc衍生基因，转染的前胸腺素α基因产生的蛋白量都是相等的。更重要的是，共转染显性负性Max基因未能降低COS细胞中内源性前胸腺素α基因或COS或L细胞中转染的野生型基因的转录。综上所述，这些数据不支持Myc激活完整人类前胸腺素α基因或模拟其结构的报告构建体转录的观点。相反，它们表明人类前胸腺素α启动子和下游元件受到缓冲，因此对调节其他基因的转录因子的瞬时波动反应微弱（如果有反应的话）。
