Computational Analysis and Modeling of Contractile Ring Assembly

描述(申请人提供)：在细胞质分裂过程中，肌动蛋白、肌球蛋白、福尔马林和相关蛋白自组装成赤道肌球蛋白收缩环。环的收缩将分裂细胞的细胞质分成两部分。虽然分子参与者的名单几乎完成，但推动环的组装和收缩的分子和集体机制仍不清楚。我们将与实验者合作开发计算工具，用于分析表达肌动蛋白(GFP-CHD)和肌球蛋白(Rlc1p-RFP)荧光标记的分裂酵母细胞的图像，这些图像揭示了有关收缩环形成模式的关键信息。然后，我们将使用这些信息来开发和测试收缩环组件的力学和动力学的数学和数值模型，并推动进一步的实验。在分裂酵母中，收缩环通过肌动蛋白依赖的一系列膜结合的“节点”的凝聚来组装，这些“节点”含有肌球蛋白Myo2p、Forin CDc12p和其他蛋白质。缩合机制是高度动态的，包括连续的肌动蛋白聚合和分解以及间歇性的节点运动。据推测，cdc12p形成肌动蛋白细丝，在被拉在一起的节点之间建立瞬时连接，并通过Myo2p马达活动形成一个环。我们将通过开发新的计算方法来测试这一假设，以分割3D静态图像中的肌动蛋白细丝，并跟踪延时电影中的肌动蛋白细丝和肌动蛋白束。这将使我们能够分析凝聚肌球蛋白网络的拓扑和动力学，并系统地量化顶点的位置、细丝长度、聚合速率、分解和捆绑。通过将这些动态与Myo2p节点的位置相关联，我们将严格地建立节点和肌动蛋白重塑位置之间的关系。对肌动蛋白运动的成功跟踪将进一步允许我们提取生物物理参数值，如细丝扩散系数、细胞质粘度和作用力。我们将模拟物理约束在节点之间建立连接的机制上的作用，以及力对Forin介导的肌动蛋白延长的影响。我们将测试环组装的全球模型，并开发统计分析和可视化方法，以便将模型预测与实验进行系统比较。细胞质分裂是细胞分裂的最后一步，由肌动蛋白和相关蛋白组成的赤道收缩环驱动。尽管破译细胞分裂的机制和控制在生物医学上具有重要意义，但收缩环组装和收缩的确切机械步骤仍不清楚。我们将通过分析细胞分裂的荧光显微镜图像，并通过建立数值和数学机制模型来解决收缩环组装的定量细节。

DESCRIPTION (provided by applicant): During cytokinesis, actin, myosin, formins and associated proteins, self-assemble into the equatorial acto-myosin contractile ring. The constriction of the ring separates the cytoplasm of dividing cells into two. While the list of molecular players is almost complete, the molecular and collective mechanisms driving the assembly and constriction of the ring remain unclear. We will collaborate with experimentalists to develop computational tools for the analysis of images of dividing fission yeast cells expressing fluorescent markers for actin (GFP-CHD) and myosin (Rlc1p-RFP), which reveal crucial information on the pattern of contractile ring formation. We will then use the information to develop and test mathematical and numerical models of the mechanics and dynamics of contractile ring assembly and to motivate further experiments. In fission yeast, the contractile ring assembles through the actin-dependent condensation of a broad band of membrane-bound "nodes" containing myosin Myo2p, formin Cdc12p, and other proteins. The condensation mechanism is highly dynamic involving continuous actin polymerization and disassembly and intermittent node motions. Cdc12p presumably nucleates actin filaments which establish transient connections between nodes that are pulled together and form a ring through Myo2p motor activity. We will test this hypothesis by developing novel computational methods to segment actin filaments in 3D static images and to track actin filaments and bundles in time-lapse movies. This will enable us to analyze the topology and dynamics of condensing actomyosin networks and systematically quantify the locations of vertices, filament lengths, rates of polymerization, disassembly, and bundling. By correlating these dynamics to the locations of Myo2p nodes we will rigorously establish the relationship between nodes and sites of actin remodeling. Successful tracking of actin motions will further allow us to extract biophysical parameter values such as filament diffusion coefficients, cytoplasmic viscosity, and forces. We will model the role of physical constraints on the mechanism of establishing connections between nodes and the effect of force on formin-mediated actin elongation. We will test global models of ring assembly and develop statistical analysis and visualization methods for systematic comparison of model predictions to experiment. Cytokinesis, the final step of cell division, is driven by the constriction of an equatorial contractile ring consisting of actin and associated proteins. Despite the biomedical importance of deciphering the mechanisms and controls of cell division, the precise mechanistic steps of contractile ring assembly and constriction remain unclear. We will resolve quantitative details of the assembly of the contractile ring by analyzing fluorescence microscopy images of dividing cells and by developing numerical and mathematical mechanistic models.