Okuno H, Akahori A, Sato H, Xanthoudakis S, Curran T, Iba H
Department of Tumor Virus Research, University of Tokyo, Japan.
Oncogene. 1993 Mar;8(3):695-701.
Fos and Jun form dimeric complexes that bind to DNA sequences containing activator protein 1 (AP-1) sites and regulate gene expression. The in vitro DNA-binding activity of these proteins is sensitive to reduction-oxidation (redox). Reduction of a single conserved cysteine residue, located in the DNA-binding domain, either by reducing agents or by a nuclear redox factor (Ref-1), is required for AP-1 DNA-binding activity. Replacing the critical cysteine with serine results in a protein that can bind to DNA in vitro even under oxidizing conditions. To determine whether redox control affects the function of Fos in vivo, we have constructed, and compared the properties of, retroviral vectors expressing either a truncated Fos protein (F118-211) or a truncated Fos protein in which the critical cysteine was replaced by serine (FC154S). In infected chicken embryo fibroblasts (CEFs), both vectors expressed similar levels of Fos protein, which formed heterodimers with Jun at equivalent efficiencies. However, extracts from cells expressing FC154S exhibited a threefold increase in AP-1 DNA-binding activity compared with cells expressing F118-211. Furthermore, this enhanced binding activity was resistant to treatment with the oxidizing agent diamide. Infection of CEFs by virus expressing FC154S resulted in increased numbers of transformed colonies and an increase in colony size compared with those obtained following infection by virus expressing Fos 118-211. These results suggest that redox regulation may limit the total level of functional Fos-Jun complexes in vivo and that escape from this control enhances transforming activity.
Fos和Jun形成二聚体复合物，该复合物可与含有活化蛋白1（AP-1）位点的DNA序列结合并调节基因表达。这些蛋白质的体外DNA结合活性对还原-氧化（氧化还原）敏感。AP-1 DNA结合活性需要通过还原剂或核氧化还原因子（Ref-1）还原位于DNA结合域中的单个保守半胱氨酸残基。将关键半胱氨酸替换为丝氨酸会产生一种即使在氧化条件下也能在体外与DNA结合的蛋白质。为了确定氧化还原控制是否在体内影响Fos的功能，我们构建并比较了表达截短的Fos蛋白（F118-211）或关键半胱氨酸被丝氨酸取代的截短Fos蛋白（FC154S）的逆转录病毒载体的特性。在感染的鸡胚成纤维细胞（CEF）中，两种载体表达的Fos蛋白水平相似，它们与Jun形成异二聚体的效率相当。然而，与表达F118-211的细胞相比，表达FC154S的细胞提取物的AP-1 DNA结合活性增加了三倍。此外，这种增强的结合活性对氧化剂二酰胺的处理具有抗性。与表达Fos 118-211的病毒感染后获得的菌落相比，表达FC154S的病毒感染CEF导致转化菌落数量增加且菌落尺寸增大。这些结果表明，氧化还原调节可能会限制体内功能性Fos-Jun复合物的总水平，而摆脱这种控制会增强转化活性。
