DOMAINS INVOLVED IN REGULATION OF RAF ACTIVITY

涉及英国皇家空军活动监管的领域

• 批准号: 3874700
• 负责人: G FANNING-HEIDECKER
• 金额: --
• 依托单位: DIVISION OF CANCER EPIDEMIOLOGY AND GENETICS
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/3874700
• 关键词: Polyomavirus; X ray crystallography; bacterial proteins; biological signal transduction; cytoplasm; enzyme structure; enzyme substrate; gene expression; genetic enhancer element; genetic promoter element; genetic transcription; growth factor receptors; human tissue; immunofluorescence technique; insect virus; membrane proteins; mitogens; molecular oncology; monoclonal antibody; mutant; neoplasm /cancer genetics; oncogenes; oncogenic virus; oncoproteins; plasmids; protein kinase; protein signal sequence; protein structure function; protein transport; serine; threonine; transfection; viral carcinogenesis

The protein products of the raf gene family are cytoplasmic serine/threonine protein kinases. They are active in the transmission of mitogenic signals from membrane-associated tyrosine kinase growth factor receptors to the nucleus. The Raf protein itself is phosphorylated and its kinase is activated by several of the tyrosine kinases. Immunofluorescence studies showed that, upon stimulation, c-Raf protein translocates from a diffuse cytoplasmic distribution to the perinuclear space, and probably to the nucleus. This assay is being used to assess the effect a series of mutations have on Raf function. Mutations in the amino-terminal half such as the cysteine array are being assayed for their response to mitogenic signals, while Raf proteins with mutations in the kinase domain are more likely to be affected in their substrate interaction. Several mutations in the carboxy-terminal half have also been incorporated into a truncated version of Raf which has been shown to be oncogenically activated. This will allow us not only to identify mutations that interfere with Raf function absolutely, but also those that render Raf thermosensitive. These mutations are hard to identify in an otherwise normal Raf against the background of constitutively expressed endogenous Raf. In addition to this in vivo assay we are also analyzing the interactions of Raf with various proteins in a aculovirus expression system. We have generated or obtained from coworkers baculoviruses which express the platelet-derived growth factor receptor and the jun and fos genes. We are currently analyzing the interactions these signal transduction components have with wild-type and mutant Raf constructs we have introduced into the baculovirus expression system. Raf protein expressed in this system as well as protein expressed in bacteria is also being used for X-ray crystallography studies and for the generation of monoclonal antibodies.

raf基因家族的蛋白产物存在于细胞质中