Mechanism of Gli3 processing in Hedgehog signaling

Hedgehog 信号传导中 Gli3 的处理机制

• 批准号: 7052092
• 负责人: BAOLIN WANG
• 金额: $ 30.27万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-05-01 至 2009-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7052092
• 关键词: DNA binding protein; biological signal transduction; biomarker; casein kinase; cell line; developmental genetics; fluorescence resonance energy transfer; laboratory mouse; phenotype; phosphorylation; proteasome; protein kinase A; protein protein interaction; protein structure function; site directed mutagenesis; transcription factor; western blottings

DESCRIPTION (provided by applicant): Cell-cell communication is a fundamental mechanism by which cell growth and differentiation are precisely controlled. Defects in these processes cause human cancer and birth defects. The Hedgehog (Hh) family of secreted signaling proteins controls cell growth and differentiation. Loss of or decrease in the Hh signaling pathway activity results in severe developmental birth defects. In addition to developmental abnormalities, inappropriate activation of the Hh signaling pathway is also associated with human cancers including basal cell carcinoma and medulloblastoma. Our long-term goal is to understand the molecular mechanisms by which Hh signal is transduced and to determine the role of Hh signaling in pattern specification and tumor formation. In Drosophila, Hh signal is mediated by Cubitus Interruptus (Ci), a zinc finger containing transcription factor. In the absence of Hh signal, a significant fraction of Ci protein is proteolytically processed to generate a transcription repressor. Hh signal stimulation blocks the Ci processing and activates the pathway. Ci processing requires its phosphorylation by PKA, CKI, and GSK3 and the activity of Slimb, a component of the novel class of ubiquitin ligase called SCF complex. Vertebrate homologs of Ci are Glil, Gli2, and Gli3. Many studies suggest that Gill and Gli2 act positively, whereas Gli3 plays a negative role in the pathway. Consistent with a negative role of Gli3 in the pathway, the majority of Gli3 protein is processed in the absence of Hh signaling. Hh signaling inhibits Gli3 processing and reduces its RNA levels, thus regulating the net output of Gli transcriptional activities. Molecular mechanisms of Gli3/Ci processing and its regulation by Hh signaling are largely unknown. Using a broadly-based approach that incorporates biochemical, cell biological, genetic, and pharmacological methods, the objectives of the proposed study are to 1) Elucidate the role of CKI and GSK3, and _TrCP, the vertebrate homolog of Slimb, in Gli3 processin.q; 2) Understand the role of the proteasome in Gli3 processin,q; 3) Determine the role of unprocessed and processed forms of Gli3 protein in vivo. The completion of the proposed study will significantly advance our understanding of the molecular mechanisms of the Hh signaling pathway. It may also shed light on the molecular mechanisms of human congenital syndromes caused by Gli3 mutations. In addition, this research may give us insights into the design of therapeutic agents that modulate Gli3 processin 9 to intervene or remedy cancer related to misregulation of the Hh signaling pathway.

描述（由申请人提供）： 细胞间通讯是精确控制细胞生长和分化的基本机制。这些过程中的缺陷会导致人类癌症和出生缺陷。Hedgehog（Hh）家族的分泌信号蛋白控制细胞生长和分化。Hh信号通路活性的丧失或降低导致严重的发育性出生缺陷。除了发育异常，Hh信号通路的不适当激活也与人类癌症相关，包括基底细胞癌和成神经管细胞瘤。我们的长期目标是了解Hh信号转导的分子机制，并确定Hh信号在模式规范和肿瘤形成中的作用。在果蝇中，Hh信号是由一种含锌指的转录因子Cubitus Interruptus（Ci）介导的。在没有Hh信号的情况下，C1蛋白的显著部分被蛋白水解加工以产生转录阻遏物。Hh信号刺激阻断Ci加工并激活通路。Ci加工需要其通过PKA、CKI和GSK 3的磷酸化以及Slim的活性，Slim是称为SCF复合物的新型泛素连接酶的组分。C1的脊椎动物同源物是Glil、Gli 2和Gli 3。许多研究表明，Gill和Gli 2发挥积极作用，而Gli 3在该途径中发挥消极作用。与Gli 3在通路中的负作用一致，大多数Gli 3蛋白在不存在Hh信号传导的情况下被加工。Hh信号抑制Gli 3加工并降低其RNA水平，从而调节Gli转录活性的净输出。Gli 3/Ci加工及其通过Hh信号调节的分子机制在很大程度上是未知的。本研究采用生物化学、细胞生物学、遗传学和药理学方法，旨在：1）阐明CKI、GSK 3和_TrCP（Slimb的脊椎动物同源物）在Gli 3加工蛋白q中的作用; 2）了解蛋白酶体在Gli 3加工蛋白q中的作用; 3）确定Gli 3蛋白未加工和加工形式在体内的作用。这项研究的完成将大大推进我们对Hh信号通路分子机制的理解。它也可能揭示Gli 3突变引起的人类先天性综合征的分子机制。此外，这项研究可能会让我们深入了解调节Gli 3加工蛋白9的治疗药物的设计，以干预或治疗与Hh信号通路失调相关的癌症。