GENETIC POLYMORPHISM IN PROSTATE CANCER

我们已经描述了NKX3.1的遗传多态性，NKX3.1是一个同源框基因， 器官特异性表达仅限于成人前列腺。 的 多态性通过用T取代C来改变密码子154，从而 导致错义密码子52用半胱氨酸取代精氨酸 （R52C）。 在对DNA样本的调查中，我们发现NKX3.1 R52 C是 存在于4%的正常捐赠者中，但在12%的前列腺捐赠者中， 癌症患者，这表明NKX3.1 R52 C可能赋予3倍的风险， 前列腺癌此外，由蛋白质编码的氨基酸改变， 多态性增加NKX3.1蛋白与DNA结合的亲合力。 因此，这种多态性代表了功能获得性突变， 似乎会影响前列腺癌的风险。 该提案侧重于DNA NKX3.1野生型的结合和转录激活， 多态性蛋白 我们将确定真正的DNA结合位点， NKX3.1。 我们将构建具有NKX3.1结合的报告质粒， 为了研究突变体和多态性之间的差异， 蛋白并进行NKX3.1的缺失分析。了建设一 嵌合NKX3.1/GAL 4融合蛋白，以研究NKX3.1 同源结构域侧翼的序列对DNA结合活性的影响， 激活已知的转录因子。 为了提高体外 表型影响的氨基酸52突变，我们产生其他氨基酸 酸取代并在报告基因测定中表征它们。 最后，通过差异显示技术，我们将识别基因 由NKX3.1调节的细胞开始理解信号传导 NKX3.1激活的通路，并研究NKX3.1应答 启动子是蛋白质的体内靶标。

We have described a genetic polymorphism of NKX3.1, a homeobox gene with organ-specific expression confined to the prostate in the adult. The polymorphism changes codon 154 by substituting a T for a C, thereby resulting in a missense codon 52 that replaces arginine with cysteine (R52C). In a survey of DNA samples we found that NKX3.1 R52C was present in 4 percent of normal donors, but in 12 percent of prostate cancer patients, suggesting that NKX3.1 R52C may confer 3-fold risk for prostate cancer. Moreover, the amino acid alteration coded by the polymorphism increases the avidity of DNA binding by the NKX3.1 protein. Therefore, this polymorphism represents a gain-of-function mutation that appears to affect prostate cancer risk. The proposal focuses on the DNA binding and transcriptional activation by NKX3.1 wild type and polymorphic proteins. We will identify the true DNA binding site for NKX3.1. We will construct reporter plasmids with the NKX3.1 binding site in order to study differences between mutant and polymorphic proteins and to perform deletion analysis of NKX3.1. We will construct chimaeric NKX3.1/GAL4 fusion proteins to study the effects of NKX3.1 sequences flanking the homeodomain on the activity of DNA binding and activation of a known transcription factor. To enhance the in vitro phenotypic impact of the amino acid 52 mutation, we generate other amino acid substitutions and characterize them in reporter gene assays. Lastly, through differential display technology, we will identify genes that are regulated by NKX3.1 to begin to understand the signaling pathways that are activated by NKX3.1 and to study NKX3.1-responsive promoters that are in vivo targets for the protein.