REGULATION OF ERCC-1 MRNA EXPRESSION BY INSULIN

工作总结：荷尔蒙和 生长因子可能控制DNA修复基因的表达 哺乳动物细胞引起了极大的兴趣，因为DNA损伤 修复可能导致基因表达异常，从而导致 退行性疾病和细胞死亡。在第一项研究中，我们 调查了ERCC1的表达是否为 核苷酸切除修复(NER)家族，受 胰岛素，如果是的话，确定了信号转导途径(S) 对胰岛素的这一作用负责。人们发现，胰岛素 ERCC1基因表达增强2~3倍 中国仓鼠卵巢(CHO)细胞过表达人胰岛素 受体和在完全分化的3T3-L1脂肪细胞中 模型以其对胰岛素的高反应性而闻名。使用各种 已知的抑制不同信号转导的药理工具 途径，我们证明了胰岛素增强ERCC1的mRNA 通过激活RAS-ERK依赖的途径而不是 磷脂酰肌醇3‘-激酶/pp70 S6激酶的参与。 DNA修复基因的表达与 细胞存活，事实证明，修复能力的缺陷 大幅提高细胞对DNA的各种损伤的敏感性。 我们发现治疗具有胰岛素样生长的CHO细胞 胰岛素样生长因子-1(IGF-1)导致ERCC1和ERCC1表达增加 XPB/ERCC3，一种编码基本基因亚基的基因产物 转录因子bTFIIH，参与NER和细胞内 周期调节。鉴于IGF-1最近一直在 作为一种抗细胞凋亡剂，我们研究了 IGF-1受体的激活可能提供对 凋亡细胞死亡是其在NER中作用的结果。细胞凋亡或 程序性细胞死亡在细胞的正常周转中起主要作用 细胞。具有特定缺陷的野生型和修复缺陷的CHO细胞 在我们的研究中使用了ERCC1或XPB/ERCC3，结果 提示IGF-1的保护作用不需要 功能NER。此外，似乎胰岛素样生长因子-1的S有能力 提供对细胞凋亡的保护依赖于 磷脂酰肌醇3‘端激活。这种可能性仍然存在 其他DNA损伤修复机制，包括错配 修复和DNA糖基酶，可能需要在介导 IGF-1的抗细胞凋亡作用。这些研究导致了 在一流同行评议期刊上发表两篇论文。

Summary of work: The notion that hormones and growth factors may control expression of DNA repair genes in mammalian cells is of great interest because impairment in DNA repair may result in aberrant gene expression leading to degenerative disorders and cell death. In the first study, we investigated whether the expression of ERCC1, a member of the nucleotide excision repair (NER) family, was under the control of insulin, and, if so, determined the signal transduction pathway(s) responsible for this action of insulin. It was found that insulin enhanced 2 to 3-fold the expression of ERCC1 mRNA both in Chinese hamster ovary (CHO) cells overexpressing human insulin receptors and in fully-differentiated 3T3-L1 adipocytes, a cell model known for its high responsiveness to insulin. Using a variety of pharmacological tools known to inhibit distinct signaling pathways, we demonstrated that insulin enhanced ERCC1 mRNA levels by activation of the Ras-ERK-dependent pathway without involvement of the phosphatidylinositol 3'-kinase/pp70 S6 kinase. There is a correlation between expression of DNA repair genes and cell survival, as evidenced by the fact that defects in repair capacity sharply increase cellular sensitivity to a variety of lesions to DNA. We have found that treatment of CHO cells with insulin-like growth factor 1 (IGF-1) led to increased expression of ERCC1 and XPB/ERCC3, a gene product that encodes a subunit of the basal transcription factor bTFIIH, which is implicated in NER and in cell cycle regulation. In light of the fact that IGF-1 has been recently identified as an antiapoptotic agent, we examined the possibility that activation of the IGF-1 receptor may offer protection against apoptotic cell death as a result of its role in NER. Apoptosis or programmed cell death plays a major role in normal turnover of cells. Wild-type and repair-deficient CHO cells with specific defects in ERCC1 or XPB/ERCC3 were used in our study, and the results indicated that the protective effect of IGF-1 did not require functional NER. Furthermore, it appears that IGF-1's ability to confer protection against apoptotic death was dependent on phosphatidylinositol 3' kinase activation. The possibility remains that other DNA lesion-repair mechanisms, which include mismatch repair and DNA glycosylases, may be required in mediating the antiapoptotic function of IGF-1. These studies have led to the publication of two papers in first-rate peer-reviewed journals.