Haneda Masataka, Xiao Hengyi, Hasegawa Tadao, Kimura Yuko, Nakashima Izumi, Isobe Ken-Ichi
Department of Basic Gerontology, National Institute for Longevity Sciences, 36-3 Gengo, Morioka-cho, Obu, Aichi 474-8522, Japan.
Gene. 2004 Jul 7;336(1):139-46. doi: 10.1016/j.gene.2004.04.008.
The GADD34 gene is transcriptionally induced by growth arrest and DNA damage. However, the mechanisms underlying the transcriptional regulation are still unclear. We analyzed the promoter of mouse GADD34 gene and the methylmethane sulfonate (MMS)-induced transcriptional regulation of this gene. By introducing genome mutants, which were linked to the luciferase reporter, into NIH3T3 cells, we defined a 100-bp fragment upstream of the transcriptional initiating site as the minimal promoter of the GADD34 gene. Subsequent study revealed that CRE-binding site located in this minimal promoter was critical for MMS-induced transcription of the GADD34 gene. In vitro binding experiments showed that phosphorylated c-Jun was contained in the CRE/DNA complex. Overexpression of the dominant negative form of c-Jun led to a decrease of MMS-responsive promoter activity. From these results, we conclude that the CRE site of the GADD34 promoter is indispensable to the MMS-responsive cis-element that c-Jun is the essential transcription factor for MMS-stimulated regulation of GADD34 gene expression and that the upstream signaling is dependent on JNK.
GADD34基因在生长停滞和DNA损伤时被转录诱导。然而，转录调控的潜在机制仍不清楚。我们分析了小鼠GADD34基因的启动子以及甲磺酸甲酯（MMS）诱导的该基因转录调控。通过将与荧光素酶报告基因相连的基因组突变体导入NIH3T3细胞，我们将转录起始位点上游100bp的片段定义为GADD34基因的最小启动子。随后的研究表明，位于该最小启动子中的CRE结合位点对MMS诱导的GADD34基因转录至关重要。体外结合实验表明，CRE/DNA复合物中含有磷酸化的c-Jun。c-Jun显性负性形式的过表达导致MMS反应性启动子活性降低。从这些结果中，我们得出结论，GADD34启动子的CRE位点对于MMS反应性顺式元件是不可或缺的，c-Jun是MMS刺激的GADD34基因表达调控的必需转录因子，并且上游信号传导依赖于JNK。
