PROTEIN KINASE C AND THYROID CELL APOPTOSIS

蛋白激酶 C 与甲状腺细胞凋亡

• 批准号: 6029543
• 负责人: JEFFREY A KNAUF
• 金额: $ 9.17万
• 依托单位: UNIVERSITY OF CINCINNATI
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-04-01 至 2002-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6029543
• 关键词: apoptosis; biological signal transduction; carcinogenesis; fusion gene; gene expression; gene mutation; genetically modified animals; laboratory mouse; p53 gene /protein; phosphorylation; posttranslational modifications; protein kinase C; thyroid gland; thyroid neoplasm

DESCRIPTION (taken from the application) In 1997, there were an estimated 16,100 new cases of thyroid cancer in the United States. Cancer is not simply a proliferation process, but the manifestation of an imbalance between cell growth and cell death. It is likely that for a tumor clone to progress the apoptotic program must be successfully disabled. In support of this paradigm, we have isolated a chimeric and truncated mutant of PKC-epsilon (Tr-PKC-epsilon), the gene for which was amplified and rearranged in a thyroid cancer cell line. When transfected into PCCL3 cells (a well-differentiated rat thyroid cell line) Tr-PKC-epsilon inhibits activation-induced translocation of the wild-type isozyme, resulting in protection of cells from apoptosis. This is accompanied by a marked impairment in p53 stabilization, which may be in part due to elevated levels of MDM2. These findings point to a role for PKC-epsilon in apoptosis signaling pathways in thyroid cells, and suggest that disruptions in PKC-epsilon function may be involved in thyroid tumorigenesis, possibly by altering the cellular response to DNA damage. In support of this we have found that in 75-85% of thyroid carcinomas there were dramatic changes in the level and/or subcellular distribution of PKC-epsilon compared to corresponding normal thyroid tissue. The following Specific Aims are proposed: (1) We will use an inducible expression system to achieve selective activation of either PKC-epsilon or the constitutively activated mutant PKC-epsilon-A159F and to determine whether this alone can initiate an apoptotic program, that can be blocked by Tr-PKC-epsilon. (2) We will explore whether PKC-epsilon activation interferes with phosphorylation, stabilization, and other post-translational modifications of p53 and MDM-2. (3) We will manipulate the function of the isozyme in thyroid follicular cells of transgenic mice, by targeting expression of either PKC-epsilon, PDK-epsilon-A159F, or the dominant negative inhibitor Tr-PKC-epsilon. Effects on thyroid cell apoptosis in vivo will then be studied in mice exposed to external radiation to the thyroid bed. (4) We will determine if the observed changes in expression and distribution of PKC-epsilon in thyroid cancers are due to somatically-acquired structural defects in the PKC-epsilon gene, or to epigenetic events. For tumor clones to expand, they must not only exhibit unrestrained stimulation to proliferate, but must also disable essential protection circuits that trigger apoptosis. We propose that PKC-epsilon is part of this defensive strategy, and that this can be subverted during tumorigenesis, or perhaps modulated during adaptive responses such as goiter involution, or thyroid remodeling.

描述（取自应用程序） 1997年，估计有16，100例甲状腺癌新病例， 美国的癌症不仅仅是一个扩散过程， 细胞生长和细胞死亡之间不平衡的表现。很可能 对于肿瘤克隆来说，凋亡程序必须成功地 禁用.为了支持这种模式，我们分离了一种嵌合的， PKC-β的截短突变体（Tr-PKC-β），其基因是 在甲状腺癌细胞系中扩增并重排。当转染到 PCCL 3细胞（一种高分化的大鼠甲状腺细胞系）Tr-PKC-β 抑制激活诱导的野生型同工酶易位， 保护细胞免于凋亡。这是伴随着一个明显的 p53稳定性受损，这可能部分是由于 MDM 2。这些发现表明PKC β在细胞凋亡信号传导中的作用 甲状腺细胞中的信号通路，并表明PKC-β功能的破坏 可能参与甲状腺肿瘤的发生，可能是通过改变细胞 DNA损伤的反应。为了支持这一点，我们发现，在75-85%的 在甲状腺癌中，存在水平和/或亚细胞的显著变化， 与相应的正常甲状腺组织相比，PKC-β的分布。 具体目的如下：（1）我们将使用一种诱导型 表达系统，以实现选择性激活PKC-β或 组成型激活的突变体PKC-BRA-A159 F，并确定这是否 单独使用可启动凋亡程序，可被Tr-PKC-β阻断。 (2)我们将探讨PKC β激活是否会干扰 磷酸化、稳定化和其他翻译后修饰 p53和MDM-2。(3)我们将操纵甲状腺同工酶的功能， 转基因小鼠的滤泡细胞，通过靶向表达 PKC-ERK、PDK-ERK-A159 F或显性负性抑制剂 Tr-PKC-β然后将研究对体内甲状腺细胞凋亡的影响 暴露于甲状腺床外部辐射的小鼠。(4)我们将确定 如果观察到的PKC-β蛋白表达和分布的变化 甲状腺癌是由于甲状腺细胞中的体细胞获得性结构缺陷， PKC-β基因，或表观遗传事件。对于肿瘤克隆的扩张， 不仅必须表现出无限制的刺激增殖，而且必须 使触发细胞凋亡的重要保护电路失效我们建议 PKC-β是这一防御战略的一部分，这是可以被颠覆的， 在肿瘤发生过程中，或者可能在适应性反应过程中调节， 甲状腺肿大退化或甲状腺重塑。