Smooth muscle molecular mechanics

平滑肌分子力学

• 批准号: RGPIN-2015-05054
• 负责人: Lauzon, AnneMarie
• 金额: $ 2.91万
• 依托单位: McGill University
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2019
• 资助国家: 加拿大
• 起止时间: 2019-01-01 至 2020-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=688040
• 关键词: Smooth; muscle; molecular; mechanics

Smooth muscle is found in all hollow organs of the body (e.g. intestine, bladder, blood vessels). Smooth muscle is unique in its ability to generate and maintain force for long periods of time, at low energy cost. Once stimulated to contract, smooth muscle can modulate its rate of energy consumption within the time course of a single contraction. This property, called the latch-state, is an important characteristic of smooth muscle but remains poorly understood. Thus, the overarching goal of this program of research is to understand the molecular mechanism behind the latch-state.*******Muscle contraction is accomplished by the interaction of the myosin molecular motors with actin filaments. It is believed that the latch-state occurs when myosin molecules get enzymatically deactivated while attached to actin. This theory came from measurements performed at the whole muscle level but was never verified at the molecular level. Several other proteins (calponin, caldesmon, non-muscle myosin, etc.) have also been hypothesized to play a role in the latch state but again, no proof of their contribution has been made at the molecular level. Thus, we aim to determine whether force can be maintained during deactivation of myosin by making force measurements directly at the molecular level. With my previous NSERC grant, we developed a micro-chamber (microfluidic device) that allows the addition of chemicals without creating any bulk flow. Thus, we now propose to use this chamber to inject deactivating enzymes during molecular force measurements and determine whether or not force maintenance (latch-state) occurs. We will also determine whether other proteins such as telokin, caldesmon etc. are needed for force maintenance or if myosin and actin are sufficient.*******Activated and non-activated myosins have very different conformations. Thus, it is possible that deactivated myosin also has its own specific temporary conformation that favours force maintenance. We propose to address this question using atomic force microscopy. Along with the molecular mechanics studies listed above, this imaging will help us get a complete picture of the latch state.*******Finally, a more recent theory of the latch-state evolved from molecular measurements that showed that partially deactivated myosin (one deactivated head out of two) interacts longer with actin than doubly- or unactivated myosin. Such myosin would remain attached to actin and maintain force. This theory has however not been verified at the whole muscle level. Thus, we propose to perform whole muscle mechanical and biochemical assays to verify whether or not such single activated heads are present during the latch state.*******In summary, this program of research proposes a multi-level (from molecule to whole muscle) and multi-disciplinary approach to understand the unique ability of smooth muscle to efficiently maintain force.******

平滑肌存在于人体所有的中空器官(如肠、膀胱、血管)中。平滑肌肉的独特之处在于，它能够以较低的能量成本长时间地产生和维持力量。一旦被刺激收缩，平滑肌就可以在一次收缩的时间过程中调节其能量消耗的速度。这一特性被称为闩锁状态，是平滑肌的一个重要特征，但人们对此知之甚少。因此，这个研究项目的首要目标是了解闭锁状态背后的分子机制。肌肉收缩是通过肌球蛋白分子马达与肌动蛋白细丝的相互作用来完成的。人们认为，当肌球蛋白分子附着在肌动蛋白上时，肌球蛋白分子被酶失活，就会发生闩锁状态。这一理论来自于在整个肌肉水平进行的测量，但从未在分子水平上得到证实。其他几种蛋白质(钙粘蛋白、钙蛋白、非肌肉肌球蛋白等)也被假设在闩锁状态中发挥作用，但同样，没有在分子水平上证明它们的贡献。因此，我们的目标是通过直接在分子水平上进行力测量来确定在肌球蛋白失活过程中是否可以保持力。在我之前的NSERC资助下，我们开发了一种微腔(微流体装置)，它允许添加化学品，而不会产生任何大流量。因此，我们现在建议使用这个小室在分子力测量期间注入失活的酶，并确定是否发生力维护(闩锁状态)。我们还将确定是否需要其他蛋白质如telokin、caldesmon等来维持力量，或者肌球蛋白和肌动蛋白是否足够。激活的和非激活的肌球蛋白具有非常不同的构象。因此，失活的肌球蛋白也可能有其特定的临时构象，这有利于力量的维持。我们建议使用原子力显微镜来解决这个问题。与上面列出的分子力学研究一起，这一成像将帮助我们对闩锁状态有一个完整的了解。*最后，从分子测量演变出的更新的闩锁状态理论表明，部分失活的肌球蛋白(两个失活的头中有一个)与肌动蛋白相互作用的时间比双倍或未激活的肌球蛋白长。这样的肌球蛋白将继续附着在肌动蛋白上，并保持力量。然而，这一理论还没有在整个肌肉水平上得到验证。因此，我们建议进行整个肌肉的机械和生化分析，以验证在闩锁状态下是否存在这样的单个激活的头部。总而言之，该研究计划提出了一个多层次(从分子到整个肌肉)和多学科的方法来了解平滑肌有效维持力量的独特能力。