IKK:Biophysical basis of dynamic regulation

描述(申请人提供)：IKK是炎症和先天免疫反应的主要信号中枢。它是一种酶复合体，接收来自大量细胞受体的信号，调节核因子-kB家族转录因子的活性。IKK的错误调节与许多慢性疾病有关，例如慢性炎症性病理(关节炎、动脉粥样硬化等)和许多不同类型的癌症。遗传证据有力地支持了IKK在许多生理和病理功能中的关键和核心作用，但它作为药物靶点的潜力尚未实现。机制和生物物理研究一直很缺乏，我们既没有对IKK调节的动力学了解，也没有结构信息。然而，有一点是明确的，IKK通过刺激特异性的严格动态控制来调节信号的特异性，并诱导特定的基因集合的表达。最近的研究报道了一种紧密耦合的激活和失活机制，该机制只能用一个多状态的激活-失活循环来描述，该循环涉及几种具有不同功能的酶的功能，如泛素连接酶、激酶、磷酸酶和折叠酶。通过对这些活动的调控，我们假设IKK周期是以刺激和细胞类型特定的方式驱动的，了解动力学关系将揭示合理定向药物干预的机会，在健康细胞中区分疾病相关的调控不当和刺激反应调节。在本方案中，我们将构建IKK循环的数学模型，以探索IKK活动的动态规律。然后，我们将重点研究特定的控制机制的生化和生物物理研究。特别是，我们测试了IKK寡聚、构象变化和上游激酶在IKK激活和抑制中的作用。计算机模拟将指导IKK动力学的遗传和药理学操作。最后，我们将重点研究肿瘤坏死因子和白介素1如何产生对IKK的差异动态控制；差异剂量反应和时间控制如何决定细胞因子陷阱的疗效。 与公共健康相关：IkappaB激酶(IKK)复合体是一种大的蛋白激酶，专门接收来自细胞表面受体的信号，并将信号传递给下游的效应转录因子NF-kB。在细胞的休眠状态下，IKK必须保持在低活性状态，并且在接收到信号时转换到高活性状态。这种严格的IKK调控模式的任何改变都会导致异常的细胞结果，包括炎症性疾病和癌症。IKK的内在和外在细胞机制如何控制IKK活性的确切机制还知之甚少。这项提案将整合包括数学建模、生物物理学、生物化学和遗传学在内的多种方法来探索IKK的调控。

DESCRIPTION (provided by applicant): IKK is the major signaling hub for inflammatory and innate immune responses. It is an enzyme complex that receives signals from a large number of cellular receptors regulates that activity of the NF-kB family of transcription factors. Misregulation of IKK is associated with many chronic diseases, such as chronic inflammatory pathologies (arthritis, atherosclerosis, etc) and many different types of cancer. Genetic evidence strongly supports IKK's critical and central role in many functions in physiology and pathology, but its potential as a drug target has not been realized. Mechanistic and biophysical studies have been lacking, and we have neither a kinetic understanding of IKK regulation nor structural information. However, what is clear is that IKK mediates signaling specificity by tight dynamic control that is stimulus-specific and induces the expression of specific sets of genes. Recent studies have reported on a tightly coupled activation and inactivation mechanism that can only be described by a multi-state activation-inactivation cycle that involves the function of several enzymes with different functions, such as ubiquitin ligases, kinases, phosphatases, and foldases. Through regulation of these activities, we hypothesize that the IKK cycle is driven in a stimulus- and cell type-specific manner, and that understanding the kinetic relationships will reveal opportunities for rationally targeted pharmacological intervention that discriminate between disease associated misregulation and stimulus-responsive regulation in healthy cells. In this proposal, we will construct a mathematical model of the IKK cycle to explore the dynamic regulation of IKK activity. We will then focus biochemical and biophysical studies on specific control mechanisms. In particular, we test the roles of IKK oligomerization, conformational changes, and upstream kinases in IKK activation and inhibition of IKK. Computational simulations will guide genetic and pharmacological manipulation of IKK dynamics. Finally, we will focus our study on how TNF and IL-1 produce differential dynamic control of IKK; how differential dose response and temporal control determine the efficacy of cytokine traps. PUBLIC HEALTH RELEVANCE: The IkappaB kinase (IKK) complex is large protein kinase that specifically receives signals from cell surface receptors and transmits the signal to downstream effector transcription factor NF-kB. IKK must remain at a low activity state under resting state of the cell and transits into a high activity state upon receiving signals. Any alteration of this tight regulatory mode of IKK results to abnormal cellular outcomes including inflammatory diseases and cancer. The precise mechanism of how IKK-intrinsic and extrinsic cellular mechanisms control IKK activity is poorly understood. This proposal will integrate diverse methodologies that include mathematical modeling, biophysics, biochemistry and genetics to probe IKK regulation.