CRCNS: Modeling Activation of CaMKII in Spines

CRCNS：模拟脊柱中 CaMKII 的激活

• 批准号: 8089566
• 负责人: MARY B KENNEDY
• 金额: $ 32.39万
• 依托单位: CALIFORNIA INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-07-01 至 2014-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8089566
• 关键词: Actins; Adverse effects; Agreement; Algorithms; Animals; Basic Science; Behavior; Binding; Biological; Biological Assay; Brain; Calcineurin; Calcium/calmodulin-dependent protein kinase; Calmodulin; Calmodulin-Binding Proteins; Catalytic Domain; Cell physiology; Chemical Engineering; Communities; Complex; Computational Technique; Computer Simulation; Cytoskeleton; Cytosol; Dendritic Spines; Development; Disease; Doctor of Philosophy; Drug Addiction; Educational process of instructing; Educational workshop; Egtazic Acid; Employment; Enzymes; Event; Excision; Female; Fire - disasters; Fostering; Foundations; Functional disorder; Glutamate Receptor; Goals; Grant; Hippocampus (Brain); Holoenzymes; Image; In Vitro; Individual; Intervention; Kinetics; Knowledge; Laboratories; Lead; Learning; Location; Long-Term Depression; Long-Term Potentiation; Medical; Membrane; Memory; Memory impairment; Mental disorders; Methods; Minority; Modeling; Molecular; Molecular Models; Multiprotein Complexes; N-Methyl-D-Aspartate Receptors; N-Methylaspartate; Names; Neuraxis; Neurobiology; Neurons; Neurosciences; Output; Pathway interactions; Phosphotransferases; Positioning Attribute; Postdoctoral Fellow; Postsynaptic Membrane; Presynaptic Terminals; Process; Property; Proteins; Publishing; Pump; Reaction; Regulation; Regulatory Pathway; Relative (related person); Research; Research Personnel; Rewards; Role; Schizophrenia; Shapes; Short-Term Memory; Signal Pathway; Signal Transduction; Signaling Protein; Staging; Structure; Students; Surface; Synapses; Synaptic Transmission; Synaptic plasticity; Tail; Techniques; Testing; Time; To specify; Training; Vertebral column; Visual; Work; base; calmodulin-dependent protein kinase II; density; dimer; graduate student; in vivo; knock-down; millisecond; molecular modeling; mutant; neural circuit; photolysis; postsynaptic; programs; protein complex; research study; scaffold; simulation; syntax

DESCRIPTION (provided by applicant): The immediate objective of this proposal is to build an accurate dynamic model of activation and autophosphorylation of the signaling protein Ca2+/calmodulin (CaM)-dependent protein kinase II (CaMKII) during influx of Ca2+ into the postsynaptic spine through NMDA receptors. The work will proceed in three stages. First, investigators will validate a model of activation of individual monomeric catalytic subunits by Ca2+ and CaM and refine its kinetic parameters by comparing the model to experiments. The deterministic model is implemented in Mathematica; the output of the model will be tested against bench assays of the enzymatic activity of monomeric subunits of CaMKII under a wide range of concentrations of the subunits, CaM, and Ca2+. The concentrations will mimic both in vivo and in vitro conditions. In the second stage, investigators will construct a model of activation of the dodecameric holoenzyme of CaMKII, based on the model validated in the first stage. They will model cooperative activation of subunit dimers within the holoenzyme, and three different paths of autophosphorylation of its subunits. The models will be constructed in the program MCell, which supports spatially correct stochastic models of protein interactions and enzymatic activation, within biologically realistic geometries. This model will employ a new rule-based algorithm to specify the locations and behavior of subunits in holoenzymes. It will be constructed in a well-mixed volume to enable testing by comparison to bench experiments with holoenzymes under a wide range of concentrations of subunits, CaM, and Ca2+. The comparisons will be used to optimize four new parameters in the holoenzyme model, and to choose the most accurate model for progression of autophosphorylation within the holoenzyme. In the third stage, investigators will introduce optimized models of the CaMKII holoenzyme into a larger MCell model of Ca2+ influx into spines through NMDA receptors and its removal by pumps and exchangers. Simulations in MCell with this model will be used to test hypotheses about parameters governing activation of CaMKII in spines. The intellectual merit of the proposal lies in its utility in the study of mechanisms of learning in the central nervous system. The regulatory machinery in a spine controls synaptic strength by regulating activity-dependent changes such as LTP and LTD. We know much about the regulatory enzymes in a spine and we have hypotheses about enzymatic networks that regulate the cellular processes controlling synaptic plasticity, including insertion and removal of glutamate receptors and changes in the shape of the spine actin cytoskeleton. However, at the present stage of analysis, qualitative studies with mutant animals, or over-expression and knock-down of particular enzymes are the dominant paradigm in the field and they are not adequate to bring our knowledge to the next level, which is to establish the timing of the action of each of these players, and the precise conditions and position in the regulatory network at which each one becomes important. To reach that level of understanding, we need better quantitative models and methods. CaMKII is one of the the initial enzymes activated by Ca2+ coming through NMDA receptors during induction of LTP. A well-validated quantitative model of its activation in the powerful MCell program will provide a starting point and an example for the construction of dynamic models of successive steps in spine regulatory pathways. The broader impacts include the educational goal of fostering introduction of computational techniques into cellular neurobiological research. A female postdoctoral fellow will be trained in experimental techniques to test computational models, and in the use of MCell. Undergraduate students (including minority students) will be involved in the work through summer research programs at Caltech and Salk. All models will be made available to the community for download. The models of CaMKII holoenzymes will be a first example of simulation in MCell of interactions within a cytosolic multiprotein complex. The syntax for doing this will be published, and taught in the regular workshops on MCell sponsored by NSF. The proposal has medical significance. Deficiencies in spine signaling pathways that use CaMKII are associated with working memory deficits similar to those that underlie schizophrenia and related thought disorders. A quantitative understanding of the factors governing activation of CaMKII during synaptic activity, and its role in controlling synaptic plasticity will facilitate development of clinically useful pharmacological agents that target specific aspects of synaptic dysfunction with fewer undesirable side effects.

描述（由申请人提供）：本提案的直接目标是建立信号蛋白 Ca2+/钙调蛋白 (CaM) 依赖性蛋白激酶 II (CaMKII) 在 Ca2+ 通过 NMDA 受体流入突触后棘过程中的激活和自磷酸化的准确动态模型。工作将分三个阶段进行。首先，研究人员将验证 Ca2+ 和 CaM 激活单个单体催化亚基的模型，并通过将模型与实验进行比较来完善其动力学参数。确定性模型在 Mathematica 中实现；该模型的输出将根据 CaMKII 单体亚基在各种亚基、CaM 和 Ca2+ 浓度下的酶活性的实验室测定进行测试。浓度将模拟体内和体外条件。在第二阶段，研究人员将基于第一阶段验证的模型构建CaMKII十二聚体全酶的激活模型。他们将模拟全酶内亚基二聚体的协同激活，以及其亚基自磷酸化的三种不同途径。这些模型将在 MCell 程序中构建，该程序支持在生物学真实几何形状内蛋白质相互作用和酶激活的空间正确随机模型。该模型将采用一种新的基于规则的算法来指定全酶中亚基的位置和行为。它将在充分混合的体积中构建，以便通过与全酶在各种亚基、CaM 和 Ca2+ 浓度下的台架实验进行比较来进行测试。这些比较将用于优化全酶模型中的四个新参数，并为全酶内的自磷酸化进展选择最准确的模型。在第三阶段，研究人员将优化的 CaMKII 全酶模型引入到更大的 MCell 模型中，该模型显示 Ca2+ 通过 NMDA 受体流入脊柱，并通过泵和交换器去除。使用该模型在 MCell 中进行的模拟将用于测试有关控制棘中 CaMKII 激活的参数的假设。 该提案的智力价值在于其在中枢神经系统学习机制研究中的实用性。脊柱中的调节机制通过调节 LTP 和 LTD 等活动依赖性变化来控制突触强度。我们对脊柱中的调节酶了解很多，并且对调节控制突触可塑性的细胞过程的酶网络有一些假设，包括谷氨酸受体的插入和去除以及脊柱肌动蛋白细胞骨架形状的变化。然而，在现阶段的分析中，突变动物的定性研究，或特定酶的过度表达和敲低是该领域的主导范式，它们不足以将我们的知识提升到一个新的水平，即确定每个参与者的行动时间，以及每个参与者在监管网络中变得重要的精确条件和位置。为了达到这种理解水平，我们需要更好的定量模型和方法。 CaMKII 是在 LTP 诱导过程中由 Ca2+ 通过 NMDA 受体激活的初始酶之一。在强大的 MCell 程序中，其激活的经过充分验证的定量模型将为构建脊柱调节途径中连续步骤的动态模型提供起点和示例。 更广泛的影响包括促进将计算技术引入细胞神经生物学研究的教育目标。一名女性博士后研究员将接受测试计算模型的实验技术以及 MCell 使用方面的培训。本科生（包括少数族裔学生）将通过加州理工学院和索尔克的暑期研究项目参与这项工作。所有模型都将提供给社区下载。 CaMKII 全酶模型将成为 MCell 中模拟胞质多蛋白复合物内相互作用的第一个例子。执行此操作的语法将被发布，并在 NSF 赞助的 MCell 定期研讨会上教授。该提议具有医学意义。使用 CaMKII 的脊柱信号通路缺陷与工作记忆缺陷相关，类似于精神分裂症和相关思维障碍的基础。对突触活动过程中控制 CaMKII 激活的因素及其在控制突触可塑性中的作用的定量理解将有助于开发临床上有用的药物，这些药物针对突触功能障碍的特定方面，且不良副作用较少。