Kinesin-2 Regulation of GLI Function in Hedgehog Signal Transduction

Kinesin-2 对 Hedgehog 信号转导中 GLI 功能的调节

• 批准号: 9274992
• 负责人: Benjamin Allen
• 金额: $ 31.75万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-06-01 至 2020-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9274992
• 关键词: Activator Appliances; Adult; Affect; Anger; Biochemical; Biochemistry; Biological Assay; Cells; Cellular biology; Chickens; Cilia; Complex; Congenital Abnormality; Data; Development; Disease; Electroporation; Embryonic Development; Equilibrium; Erinaceidae; Family; Future Generations; GLI Family Protein; GLI gene; Genetic; Genetic Transcription; Goals; Homeostasis; Image; In Vitro; Kinesin; Malignant Neoplasms; Mediating; Modeling; Motor; Mus; Natural regeneration; Organ; Organelles; Pathology; Pathway interactions; Pattern; Proteins; Regulation; Research; Resolution; Role; Signal Transduction; Testing; Therapeutic Uses; Tissues; Transcription Coactivator; Transcription Repressor/Corepressor; Transcriptional Activation; Work; anterograde transport; cellular imaging; ciliopathy; developmental genetics; gene repression; genetic approach; hedgehog signal transduction; in vivo; insight; member; novel; novel therapeutics; progenitor; protein function; protein transport; public health relevance; response; smoothened signaling pathway; trafficking; transcription factor

 DESCRIPTION (provided by applicant): The Hedgehog (HH) signaling pathway is vital for tissue patterning and organ formation during embryogenesis, as well as adult tissue homeostasis, renewal and regeneration. In contrast, aberrant HH pathway function results in numerous developmental diseases and birth defects, and is responsible for a growing number of cancers. GLI proteins (GLI1-3) are essential downstream transcriptional effectors of the HH pathway, with both transcriptional activator and repressor functions. Recent work indicates that primary cilia are essential regulators of HH pathway activity, in part through the ciliary targetin and trafficking of GLI proteins. There are critical gaps, however, in our understanding of the mechanisms by which these proteins traffic through cilia, and the consequences that this trafficking has on GLI protein function downstream of HH pathway activation. Here we propose to answer the following questions: what regulates the ciliary targeting and trafficking of GLI proteins, and how does this impact GLI transcriptional activity? The long-term goal of this research is to understand how ciliary transport of HH pathway components affects their function during HH signal transduction in development and disease. The objective of this proposal is to precisely define the mechanisms by which the GLI proteins traffic through cilia and the functional consequences of this transport on GLI-mediated HH signaling. Our data indicate that GLI proteins selectively interact with members of the kinesin-2 family of motor proteins, and that disrupting these interactions significantly impacts GLI protein function. Thus, we hypothesize that the kinesin-associated protein, KAP3, as well as the kinesin-2 motors, KIF3A, KIF3B and KIF17, are essential for ciliary trafficking of GLI proteins, and that this transport is necessary or proper GLI processing and function. The rationale for this proposal is that a deeper mechanistic understanding of the ciliary trafficking of GLI proteins will significantly inform the development f novel therapies for a growing number of ciliopathies and HH-driven pathologies where de-regulated HH signaling promotes disease initiation and progression. To test our hypothesis, we propose two specific aims. In Aim 1 we will: 1) define the physical interactions of GLI proteins with the heterotrimeric kinesin-2 motor complex, 2) determine the role of heterotrimeric kinesin-2 motors in regulating the subcellular localization and processing of GLI proteins, and 3) investigate the functional consequences of disrupting KIF3-KAP3-GLI interactions on HH signal transduction. In Aim 2 we will: 1) define novel interactions between GLI proteins and the homodimeric kinesin-2 motor, KIF17, 2) elucidate the consequences of disrupting GLI-KIF17 interactions on GLI protein localization, processing and activity, and 3) investigate a novel role for KIF17 in GLI-mediated cerebellar progenitor proliferation in vivo. The proposed research will utilize a combination of in vitro (biochemical, imaging, and cell signaling assays) and in vivo (mouse genetics and chicken in ovo electroporations) approaches to define how kinesin-2 interactions with GLI proteins affects HH pathway function. This work will define new mechanisms that regulate HH pathway function, and will significantly impact our understanding of HH-driven developmental diseases and cancers.

 描述（由适用提供）：刺猬（HH）信号通路对于胚胎发生过程中的组织模式和器官形成以及成人组织稳态，更新和再生至关重要。相比之下，异常的HH途径功能会导致许多发育疾病和先天缺陷，并导致越来越多的癌症。 GLI蛋白（GLI1-3）是HH途径的必不可少的下游转录效应，具有转录激活因子和反射器功能。最近的工作表明，原发性纤毛是HH途径活性的重要调节剂，部分是通过纤毛靶标和GLI蛋白的运输。但是，在我们对这些蛋白质通过纤毛流量的机制以及该流量对HH途径激活下游的Gli蛋白功能的后果的理解时，存在关键的差距。在这里，我们建议回答以下问题：是什么调节了GLI蛋白的纤毛靶向和运输，这如何影响GLI转录活性？这项研究的长期目标是了解HH途径成分的睫状运输如何影响其在发育和疾病中HH信号传递期间的功能。该提案的目的是精确定义Gli蛋白通过CILIA的流量以及该传输对GLI介导的HH信号的功能后果的机制。我们的数据表明，Gli蛋白选择性地与运动蛋白的驱动蛋白-2家族的成员相互作用，并且破坏了这些相互作用会显着影响Gli蛋白功能。这就是我们假设，运动蛋白相关的蛋白KAP3以及驱动蛋白-2电动机KIF3A，KIF3B和KIF17对于GLI蛋白的纤毛运输至关重要，并且该转运是必要的或必要的或适当的GLI处理和功能。该提议的理由是，对GLI蛋白的睫状流量的更深入的机械理解将显着为越来越多的纤毛病变和HH驱动的病理学的新型疗法提供了大量的信息，在这些疗法中，HH信号降低了HH信号促进疾病的起始和进展。为了检验我们的假设，我们提出了两个具体目标。 In Aim 1 we will: 1) define the physical interactions of GLI proteins With the heterotrimeric kinesin-2 motor complex, 2) determine the role of heterotrimeric kinesin-2 motors in determining the subcellular localization and processing of GLI proteins, and 3) investigate the functional consequences of disrupting KIF3-KAP3-GLI interactions on HH signal transduction.在目标2中，我们将：1）定义GLI蛋白与同型二聚体运动蛋白-2电动机之间的新型相互作用，KIF17，2）阐明了破坏Gli蛋白定位，加工和活性以及3）在Gli介导的脑介导的脑脑中的新作用的GLI KIF17相互作用的后果。拟议的研究将利用体外（生化，成像和细胞信号测定）和体内（OVO电孔中的小鼠遗传学和鸡肉）的组合来定义驱动蛋白-2与Gli蛋白与Gli蛋白的相互作用如何影响HH途径功能。这项工作将定义调节HH途径功能的新机制，并会严重影响我们对HH驱动的发育疾病和癌症的理解。