The mechanical control of mechanosensation

 描述(申请人提供)：机械力在对触摸、声音和光的反应中起着关键作用，但也对我们身体产生的压力起着关键作用。我们血管系统中的特殊感觉细胞会随着我们心脏的每一次跳动而周期性地变形，但仍然足够敏感，可以监测我们身体的机械状态。我们动脉中的这些压力感受器和肺部的伸展感受器的感觉能力的破坏可能会导致心血管和肺部疾病。尽管这对我们的生理学很重要，但由于研究机械力对单个细胞的影响的挑战，导致机械力感觉的因素仍然难以捉摸。虽然测量单个细胞并对其施加力的技术正在变得可行，但检测活细胞内这种变形的工具还不存在。为了研究神经元中的基本机械转导途径，这一建议在遗传模式生物线虫的背景下整合了物理学、生物学和工程学。在线虫的302个神经元中，有60个是由机械力激活的。特别是触摸感受器神经元，在其已知的生理反应和分子机制方面具有非常好的特征，因此是研究细胞对机械应激反应的强大系统。此外，线虫的神经元在动物运动时会受到周期性变形，这是由于正弦运动模式，类似于我们身体中的本体感知机械传感器。重要的是，机械-电转导(MET)的许多组成部分在哺乳动物中是保守的。这个项目的指导阶段将确定在触摸和本体感觉过程中导致MET通道MEC-2开放的生化力传递途径，MEC-2是一种在哺乳动物对机械力的反应中至关重要的保守蛋白质。Aim 1将结合设计和实施一种新的设备来可视化活蠕虫中MEC-2的机械力效应，并将利用经典的生物化学和遗传筛选来定义MEC-2与细胞细胞骨架组件的相互作用。这些结果将第一次展示强制门是如何建立真核细胞的MET通道的，这与躯体感觉神经科学和听力领域的广泛研究社区有关。在长期、独立的阶段(目标3)，该项目将致力于了解分子和细胞的机械特性如何影响自由行为的动物的决策，并在分子、细胞和系统水平上解决机械感觉的机制。这将包括通过神经元激活的钙成像表征线虫中拉伸感受器的本体感受器，弹性底物上分离的本体感受器培养的靶向刺激，以及携带修饰细胞骨架蛋白的转基因蠕虫的运动行为。这些实验将是第一次将分子力学与触摸和运动等特定行为表型联系起来。

 DESCRIPTION (provided by applicant): Mechanical force plays pivotal roles in the responses to touch, sound and light, but also to stresses generated in our own body. Specialized sensory cells in our vascular system are periodically deformed with every beat of our heart, yet remain sensitive enough to monitor the mechanical state of our body. Disruption of the sensory capacity of these baroreceptors in our arteries and stretch receptive proprioceptors in our lungs can lead to cardiovascular and pulmonary diseases. Despite this importance to our physiology, the factors contributing to the sense of mechanical force remain elusive due to challenges in studying their consequences on individual cells. Whereas the technology to measure and exert forces on single cells is becoming available, the tools to detect such deformations inside living cells do not yet exist. To investigate the basic mechanotransduction pathways in neurons, this proposal integrates physics, biology, and engineering within the context of the genetic model organism, C. elegans. Of the 302 neurons in C. elegans, 60 are activated by mechanical force. Touch receptor neurons, in particular, are extremely well characterized in terms of their known physiological responses and molecular machinery, and are thus a powerful system to study the cellular response to mechanical stress. Moreover, C. elegans neurons are subjected to cyclic deformation as the animal moves due to the sinusoidal locomotion pattern, analogous to proprioceptive mechanosensors in our own bodies. Importantly, many components of mechano-electrical transduction (MeT) are conserved in mammals. The mentored phase of this project will identify the biochemical force transmission pathway during touch and proprioception that leads to the opening of the MeT channel MEC-2, a conserved protein pivotal to the response to mechanical force in mammals. Aim 1 will utilize a fluorescent force reporter assay in conjunction with the design and implementation of a novel device to visualize mechanical force effects on MEC-2 in live worms, and Aim 2 will define the interaction of MEC-2 with components of the cellular cytoskeleton using classical biochemistry and genetic screens. These results will show, for the first time, how force gates a eukaryotic MeT channel, relevant to a wide research community in somatosensory neuroscience and hearing. In the long-term, independent phase (Aim 3), this project will aim to understand how mechanical properties of molecules and cells influence decisions of a freely behaving animal and resolve the mechanism of mechanosensation on the molecular, cellular, and systems level. This will involve the characterization of stretch-receptive proprioceptors in C. elegans by calcium imaging of neuron activation, targeted stimulation of isolated proprioceptor culture on elastic substrates and locomotive behavior of transgenic worms carrying modified cytoskeletal proteins. These experiments will be the first that link molecular mechanics to specific behavioral phenotypes such as touch and locomotion.