Mouse p53 represses the rat brain creatine kinase gene but activates the rat muscle creatine kinase gene
Mouse p53 represses the rat brain creatine kinase gene but activates the rat muscle creatine kinase gene
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DOI：
10.1128/mcb.14.12.8483-8492.1994
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发表时间：
1994-12
影响因子：
5.3
通讯作者：
Jiuqiao Zhao;Schmieg Fi;D. Simmons;G. Molloy
中科院分区：
文献类型：
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作者：
Jiuqiao Zhao;Schmieg Fi;D. Simmons;G. Molloy
肌酸激酶 (CK) 为高能量消耗的细胞反应再生 ATP。虽然肌肉 CK (CKM) 几乎只在成人骨骼肌和心肌中表达，但脑 CK (CKB) 表达更广泛，并且在脑胶质细胞中表达最高。 CKB 在人肺肿瘤细胞中表达也很高，其中许多细胞含有 p53 等位基因突变。我们最近在 HeLa 细胞中检测到高水平的 CKB mRNA，并在这项研究中测试了这是否可能是由于 HeLa 细胞中 p53 蛋白含量极低所致。瞬时转染实验表明，野生型小鼠 p53 在 HeLa 细胞中严重抑制大鼠 CKB 启动子，但在 CV-1 猴肾细胞中则不然，这表明，在 HeLa 细胞而非 CV-1 细胞中，p53 要么与所需的辅阻遏物相关，要么经历 CKB 抑制所需的翻译后修饰。相反，小鼠野生型 p53 在 CV-1 细胞中强烈激活大鼠 CKM 启动子，但在 HeLa 细胞中则不然，这表明，在 CV-1 细胞中，p53 可能与所需的共激活因子结合或以 CKM 激活所需的方式进行修饰。发现 p53 介导的调节所需的 DNA 序列位于 CKB 启动子的 -195 至 +5 bp 内和 CKM 启动子的 -168 至 -97 bp 内。此外，来自近端大鼠 CKM 启动子的 112 bp 片段（bp -168 至 -57）包含五个简并 p53 结合元件，能够在 CV-1 细胞中的异源启动子上赋予 p53 介导的激活。而且，当这种新的p53序列位于天然168-bp大鼠CKM启动子中时，赋予p53介导的激活等于或大于最初表征的远上游(bp -3160)小鼠CKM p53元件的激活。因此，CKB和CKM可能是少数可以作为体内p53靶标的细胞基因之一。此外，我们分析了一系列 p53 保守区 II 发生改变的错义突变体。突变对 p53 反式抑制和反式激活活性的影响不同，表明 p53 中的这些活性是可分离的。 p53突变体反式激活的能力与其抑制腺病毒E1a和激活的Ras对大鼠胚胎成纤维细胞转化的能力密切相关。
The creatine kinases (CK) regenerate ATP for cellular reactions with a high energy expenditure. While muscle CK (CKM) is expressed almost exclusively in adult skeletal and cardiac muscle, brain CK (CKB) expression is more widespread and is highest in brain glial cells. CKB expression is also high in human lung tumor cells, many of which contain mutations in p53 alleles. We have recently detected high levels of CKB mRNA in HeLa cells and, in this study, have tested whether this may be due to the extremely low amounts of p53 protein present in HeLa cells. Transient transfection experiments showed that wild-type mouse p53 severely repressed the rat CKB promoter in HeLa but not CV-1 monkey kidney cells, suggesting that, in HeLa but not CV-1 cells, p53 either associates with a required corepressor or undergoes a posttranslational modification necessary for CKB repression. Conversely, mouse wild-type p53 strongly activated the rat CKM promoter in CV-1 cells but not in HeLa cells, suggesting that, in CV-1 cells, p53 may associate with a required coactivator or is modified in a manner necessary for CKM activation. The DNA sequences required for p53-mediated modulations were found to be within bp -195 to +5 of the CKB promoter and within bp -168 to -97 of the CKM promoter. Moreover, a 112-bp fragment from the proximal rat CKM promoter (bp -168 to -57), which contained five degenerate p53-binding elements, was capable of conferring p53-mediated activation on a heterologous promoter in CV-1 cells. Also, this novel p53 sequence, when situated in the native 168-bp rat CKM promoter, conferred p53-mediated activation equal to or greater than that of the originally characterized far-upstream (bp -3160) mouse CKM p53 element. Therefore, CKB and CKM may be among the few cellular genes which could be targets of p53 in vivo. In addition, we analyzed a series of missense mutants with alterations in conserved region II of p53. Mutations affected p53 transrepression and transactivation activities differently, indicating that these activities in p53 are separable. The ability of p53 mutants to transactivate correlated well with their ability to inhibit transformation of rat embryonic fibroblasts by adenovirus E1a and activated Ras.