STUDIES OF THE FOS ONCOGENE

癌基因的发现彻底改变了对癌症分子基础的分析。 癌基因为从分子水平上剖析控制正常细胞生长的信号通路和阐明导致肿瘤发生的事件提供了起点。 fos癌基因负责通过Finkel-Biskis-Jinkins鼠肉瘤病毒诱导骨肉瘤和细胞转化。 其蛋白产物Fos作为可诱导的哺乳动物转录因子AP 1的组分起作用。 c-fos表达的诱导与神经元的有丝分裂、分化、凋亡和兴奋有关。 它被认为参与复杂的信号转导网络，通过调节基因表达将细胞外刺激与长期细胞反应偶联。fos基因的转化被认为是基因表达调控不当的结果。 在本申请中提出了三个主要的具体目标。 第一个问题是通过代表性差异分析和cDNA阵列杂交方法鉴定和表征负责细胞转化的fos靶基因。 在第二个具体的目标，我们将跟进的初步研究表明，DNA 5-甲基胞嘧啶转移酶-1（dnmt 1）是一个参与细胞转化的fos靶基因。 我们计划在fos转化的细胞中鉴定被dnmt 1甲基化的基因，并分析它们对转化的贡献。 第三个具体目标涉及还原/氧化（氧化还原）在Fos功能和细胞生长调节中的作用。以前，我们证明了双功能氧化还原/核酸内切酶基因，ref-1，通过DNA结合结构域中的保守半胱氨酸残基调节Fos的DNA结合活性。 小鼠Ref-1基因的破坏导致早期胚胎死亡。 我们将通过胚胎干细胞中的同源重组将点突变引入Ref-1，所述同源重组独立地消除Ref-1的氧化还原和核酸酶功能。 这些细胞将用于产生突变小鼠和细胞系，这些突变小鼠和细胞系将用于分析Ref-1在调节AP 1活性和细胞生长中的作用。

The discovery of oncogenes revolutionized analysis of the molecular bases of cancer. Oncogenes provided starting points for molecular dissection of the signaling pathways that govern normal cell growth and for elucidation of the events that lead to oncogenesis. The fos oncogene is responsible for induction of osteosarcomas and cell transformation by the Finkel-Biskis-Jinkins murine sarcoma virus. Its protein product, Fos, functions as a component of the inducible mammalian transcription factor AP 1. Induction of c-fos expression is associated with mitogenesis, differentiation, apoptosis and excitation of neurons. It is believed to participate in complex signal transduction networks that couple extracellular stimuli to long-term cellular responses by regulating gene expression. Transformation by fos is thought to be a consequence of inappropriate regulation of gene expression. Three major specific aims are addressed in this application. The first concerns the identification and characterization of fos target genes responsible for cell transformation by Representational Difference Analysis and cDNA array hybridization approaches. In the second specific aim we will follow up preliminary studies demonstrating that DNA 5-methylcytosine transferase-1 (dnmt1) is a fos target gene involved in cell transformation. We plan to identify genes that are methylated by dnmt1 in fos transformed cells and analyze their contribution to transformation. The third specific aim concerns the role of reduction/oxidation (redox) in the regulation of Fos function and cell growth. Previously, we demonstrated that the bifunctional redox/endonuclease gene, ref-1, regulates the DNA binding activity of Fos through a conserved cysteine residue in the DNA binding domain. Disruption of the Ref-1 gene in mice causes early embryonic lethality. We will introduce point mutations into Ref-1, by homologous recombination in embryonic stem cells that ablate the redox and nuclease functions of Ref-1 independently. These cells will be used to generate mutant mice and cell lines that will be used to analyze the role of Ref-1 in modulating AP1 activity and cell growth.