Mechanotransduction via LIM Domain Protein Mechanosensing

通过 LIM 结构域蛋白机械传感进行机械转导

基本信息

批准号：
10735689
负责人：
Patrick William Oakes
金额：
$ 30.68万
依托单位：
LOYOLA UNIVERSITY CHICAGO
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-09-01 至 2028-07-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10735689
关键词：
Ablation Actins Adhesions Affect Architecture Binding Binding Proteins Binding Sites Biochemical Breathing Cancerous Cells Cellular Structures Cellular biology Chimera organism Cytoskeleton Data Development Disease Environment Extracellular Matrix Family Family member Focal Adhesions Impairment Individual Integrins Kinetics Knowledge LIM Domain LIM Domain Protein Lasers Location Machine Learning Measures Mechanics Mediating Microfilaments Molecular Conformation Motor PTK2 gene Phenylalanine Phosphorylation Physiological Processes Physiology Play Polymers Process Proteins Research Role Signal Pathway Signal Transduction Signaling Protein Site Smooth Muscle Myocytes Stress Stress Fibers Stretching Structure Talin Testing Tissues Traction Vinculin Work ZYX gene adhesion receptor biophysical techniques cell behavior experimental study extracellular insight mechanical force mechanical signal mechanotransduction migration new therapeutic target novel optogenetics paxillin polymerization pressure protein purification recruit repaired respiratory smooth muscle response transmission process

项目摘要

Project Summary Mechanical interactions play a fundamental role in physiology, allowing cells to move, generate forces, and assemble into multicellular structures. Key to these processes is the ability of cells to turn mechanical signals into biochemical signals, an activity known as mechanotransduction. The search for mechanosensitive proteins that could facilitate mechanotransduction has primarily focused on proteins that undergo conformational changes in response to force or proteins that display changes in the bond kinetics under load. There exists another class of proteins, however, that recognize structures under strain. The canonical example of this class of proteins is the LIM domain protein zyxin, which recognizes strained actin stress fibers and recruits actin polymerization factors to repair them. Recent work has highlighted that the strain sensing mechanism of the LIM domains is not unique to zyxin and that numerous other members of the family of LIM domain proteins display a similar ability. This suggests LIM domain proteins could act as mechanotransducers, recognizing strain via their LIM domains and converting it to other biochemical signals via interactions with the other domains in the protein. To explore this hypothesis further it is crucial that we understand how LIM domains recognize strained actin filaments, and how those interactions propagate signals downstream of the strain sites. Here we propose to establish rigorous experimental strategies to decipher the mechanisms underlying LIM domain protein mechanotransduction. We employ a combination of biophysical techniques including laser ablations, optogenetics and cell stretching to quantitatively and repeatedly induce strain sites in the actin cytoskeleton. In Aim 1 we will test alternative mechanisms of LIM domain strain sensing by comparing proteins from the testin family of LIM domain proteins which require only a single LIM domain to recognize strain sites, compared to the three tandem LIM domains that zyxin requires. In Aim 2 we will test whether in addition to stretched actin filaments in stress fibers, LIM domain proteins can recognize other strained actin structures, such as compressed stress fibers or actin meshworks. Finally, in Aim 3 we will investigate how binding of LIM domain proteins to strain sites leads to a propagation of that mechanical signal to other parts of the stress fiber and the extracellular matrix. Together these studies will greatly expand our knowledge of mechanotransduction and provide insight into this fundamental signaling mechanism.

项目概要机械相互作用在生理学中发挥着基础作用，允许细胞移动、产生力和组装成多细胞结构。这些过程的关键是细胞转换机械信号的能力转化为生化信号，这种活动称为机械转导。寻找机械敏感蛋白可以促进机械转导的研究主要集中在经历构象的蛋白质对力或蛋白质的响应发生变化，在负载下显示出键动力学的变化。存在然而，另一类蛋白质可以识别应变下的结构。这类蛋白质的典型例子是 LIM 结构域蛋白 zyxin，它识别应变肌动蛋白应激纤维并招募肌动蛋白聚合因子来修复它们。最近的工作强调了 LIM 结构域的应变传感机制并非 zyxin 所独有，而且 LIM 结构域的许多其他成员 LIM 结构域蛋白家族也表现出类似的能力。这表明 LIM 结构域蛋白可以充当机械传感器，通过其 LIM 结构域识别应变并将其转换为其他生化信号通过与蛋白质中其他结构域的相互作用。为了进一步探索这个假设，至关重要的是我们了解 LIM 结构域如何识别应变肌动蛋白丝，以及这些相互作用如何传播信号应变位点的下游。在这里，我们建议建立严格的实验策略来破译 LIM 的机制结构域蛋白机械转导。我们采用包括激光在内的生物物理技术组合消融、光遗传学和细胞拉伸以定量且重复地诱导肌动蛋白中的应变位点细胞骨架。在目标 1 中，我们将通过比较蛋白质来测试 LIM 域应变传感的替代机制来自 LIM 结构域蛋白的 testin 家族，仅需要一个 LIM 结构域即可识别菌株位点，与 zyxin 所需的三个串联 LIM 结构域相比。在目标 2 中，我们将测试是否除了应力纤维中拉伸的肌动蛋白丝，LIM 结构域蛋白可以识别其他应变的肌动蛋白结构，例如压缩应力纤维或肌动蛋白网状结构。最后，在目标 3 中，我们将研究 LIM 的结合如何应变位点的结构域蛋白导致机械信号传播到应力纤维的其他部分和细胞外基质。这些研究将极大地扩展我们对力转导的了解并提供对这一基本信号机制的深入了解。