Role of NF2 tumor suppressor gene promoter in the tumorigenesis

NF2抑癌基因启动子在肿瘤发生中的作用

• 批准号: 11671371
• 负责人: TAKESHIMA Hideo
• 金额: $ 2.3万
• 依托单位: Kagoshima University (2000)Kumamoto University (1999)
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1999
• 资助国家: 日本
• 起止时间: 1999 至 2000
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11671371/
• 关键词: NF2; gene; promoter; cloning; luciferase; methylation

Background : Although mutational inactivation and allelic loss in the NF2 gene appear to be causal events in the majority of vestibular schwannomas, involvement of another potentially important mechanism, transcriptional inactivation, has not been investigated. Results : We cloned and functionally characterized the 5'-flanking region of the human NF2 gene and identified the molecular mechanisms that regulate NF2 expression. Luciferase assay and site-directed mutagenesis demonstrated that a 70-base pair (bp) region (-591 to -522 bp from the translation start site) was essential for the basic expression of the NF2 gene. Gel mobility shift assay indicated recognition by nuclear protein(s) of the unusually long (〜66 bp) sequences in this region. Recognition was inhibited by either mutation of the binding core sequence or by methylation of three CpG sites. Point mutations at these CpG sites significantly decreased promoter activity, suggesting the importance of these sites. In 14 of 23 vestibular schwannomas, these 3 CpG sites within this region were methylated in a site-specific manner and the methylation status was consistent with the expression of NF2 mRNA.Conclusions : Suppressed expression by aberrant methylation or mutation of the promoter elements could be an alternative mechanism for inactivation of the NF2 gene.

背景：虽然NF2基因的突变失活和等位基因丢失似乎是大多数前庭神经鞘瘤的原因，但另一个潜在的重要机制，即转录失活，尚未被研究。结果：我们克隆了人NF2基因的5‘侧翼区，并对其进行了功能鉴定，确定了调控NF2表达的分子机制。荧光素酶分析和定点突变表明，NF2基因的一个70个碱基对的区域(从翻译起始点开始-591到-522个碱基对)是NF2基因基本表达所必需的。凝胶迁移率改变分析表明，核蛋白(S)识别该区域的异常长(~66bp)序列。识别被结合核心序列的突变或三个CpG位点的甲基化所抑制。这些CpG位点的点突变显著降低了启动子的活性，这表明了这些位点的重要性。在23例前庭神经鞘瘤中，14例该区域的3个CpG位点甲基化状态与NF2基因的表达一致。结论：启动子元件的异常甲基化或突变抑制了NF2基因的表达，可能是NF2基因失活的另一种机制。

项目成果

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Kino, T.、Takeshima, H.、Nishi, T.、Yamamoto, K.、Nakao, M.、Saito, Y.、Kochi, M.、Kuratsu, J.、Saya, H. 和 Ushio, Y.。

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