Polycomb repressor complex protein EZH2 regulation of muscle stem cell migration and differentiation

由于运动、年龄、疾病或不良生活方式，肌肉在一生中不断受损。但与成人身体中的许多组织不同，肌肉能够通过位于肌肉纤维边缘的特殊肌肉干细胞的作用进行自我修复。当它们检测到损伤时，肌肉干细胞会移动到损伤部位，然后整合到受损的纤维中，这一过程被称为分化。肌肉无力，例如在我们变老或癌症等疾病期间，被认为是由于肌肉干细胞无法修复受损的肌肉，而是连接细胞和脂肪细胞取代了肌肉纤维。更好地了解肌肉干细胞移动到损伤部位和分化的方式，最终将有助于开发预防或治疗肌肉虚弱的疗法。作为对损伤的反应，肌肉干细胞也会打开或关闭特定的基因，使用DNA中的控制区，称为表观遗传开关。这项研究旨在研究一种名为EZH2的酶，它可以改变DNA中的这些表观遗传开关。因此，EZH2可以潜在地打开或关闭正确的基因，以响应肌肉损伤，从而导致肌肉修复。事实上，我们知道EZH2会影响肌肉修复，因为当这种酶在小鼠的肌肉中不起作用时，肌肉就没有得到适当的修复。但当肌肉干细胞修复受损的肌肉时，我们并不确切地知道EZH2控制了哪些开关，从而控制了哪些基因。这项研究的目的之一是识别这些基因。我们现在有了令人兴奋的初步结果，表明EZH2不仅影响分化，而且还影响肌肉干细胞向损伤部位的移动。现在，我们希望了解EZH2如何控制肌肉干细胞向损伤方向的迁移以及它们在损伤部位的分化。为了做到这一点，我们将使用斑马鱼的幼虫，因为斑马鱼的肌肉再生速度很快，而且幼虫是透明的，所以我们可以很容易地用显微镜看到肌肉干细胞的运动。我们将使用带有荧光标记的肌肉干细胞的斑马鱼幼体来测量肌肉干细胞是如何向损伤方向移动的。鱼体内的荧光细胞将被强大的显微镜观察，以制作4D电影，并将使用专门开发的计算机软件来跟踪细胞。这不是在哺乳动物等其他不透明的动物身上可以轻易做到的事情。斑马鱼的肌肉修复也以与人类大致相同的方式进行，相似的基因在损伤后被打开和关闭，这使这成为研究肌肉修复过程的一个很好的系统。我们将测量当EZH2被阻止工作时，肌肉干细胞在受伤后的表现。然后我们会问EZH2在修复过程中的哪个点是重要的，因为我们可以直观地看到在损伤之前、期间和之后肌肉干细胞中哪些基因被打开或关闭。为了证明EZH2是否通过表观遗传开关控制基因，我们将提取迁徙的肌肉干细胞，并测量DNA上的开关。对癌细胞的研究表明，EZH2还可以单独控制细胞的运动，而不是调节基因的开启或关闭。我们还将研究EZH2是否在肌肉干细胞中也具有这种活性，方法是使用在癌细胞中发现的EZH2的突变版本，并询问它们是否改变了肌肉干细胞的移动方式。我们的结果对于理解“表观遗传”开关如何与肌肉干细胞移动到受损部位以确保肌肉修复成功将是重要的。我们还将能够识别在肌肉修复过程中由EZH2控制的基因，因此可能能够针对这些基因作为提高肌肉力量的治疗方法。

Muscle is constantly damaged throughout life, due to exercise, age, sickness or poor lifestyle. But unlike many tissues in the adult body, muscle is able to repair itself through the action of specialised muscle stem cells which lie at the edges of muscle fibres. When they detect damage muscle stem cells move to the site of damage and then integrate into the damaged fibres, in a process called differentiation. Muscle weakness, for instance as we age or during disease such as cancer, is thought to occur due to an inability of muscle stem cells to repair damaged muscle, and instead connective and fat cells take the place of muscle fibres. A better understanding of the ways muscle stem cells move to the site of damage and differentiate will ultimately allow the development of therapies to prevent or treat muscle weakness.In response to damage, muscle stem cells also turn specific genes on or off, using control regions in the DNA called 'epigenetic' switches. This study aims to investigate an enzyme, EZH2, that makes changes to these epigenetic switches in the DNA. EZH2 can therefore potentially turn the correct genes on or off in response to muscle damage and so lead to repair of the muscle. Indeed we know that EZH2 can influence muscle repair, because when this enzyme does not work in the muscle of mice, the muscle is not repaired properly. But we do not know exactly which switches, and therefore genes, are controlled by EZH2 when muscle stem cells repair damaged muscle. One aim of this study is to identify those genes. We now have exciting preliminary results that reveal EZH2 not only influences differentiation, but also influences muscle stem cell movement towards sites of damage. We now wish to understand how EZH2 can control both migration of muscle stem cells towards damage and their differentiation at the site of damage. To do this we will use zebrafish larvae, because the muscle of zebrafish regenerates quickly, and the larvae are transparent so that we can easily see the movement of muscle stem cells using a microscope. We will measure how muscle stem cells move towards injury by using zebrafish larvae that have fluorescently labelled muscle stem cells. Fluorescent cells in the fish will be observed by powerful microscopes to make 4D movies and an especially developed computer software will be used to track cells. This is not something that can easily be done in other, non-transparent, animals such as mammals. Muscle repair in zebrafish also occurs in much the same way as in humans and similar genes are turned on and off in response to damage, making this a good system to study muscle repair processes.We will measure how muscle stem cells behave after injury when EZH2 is prevented from working. We will then ask at which point in the repair process EZH2 is important, as we can visualise which genes are turned on or off in muscle stem cells before, during and after injury. To show if EZH2 controls genes by epigenetic switches we will extract migrating muscle stem cells and measure the switches on the DNA. Studies of cancer cells suggest that EZH2 can also control the movement of cells separately from regulating how genes are turned on or off. We will also investigate whether EZH2 has this activity in muscle stem cells as well, by using mutated versions of EZH2 found in cancer cells and asking if they change the way that the muscle stem cells move. Our results will be important for understanding how the 'epigenetic' switches are coordinated with the movement of muscle stem cells to sites of damage to ensure that muscle repair is successful. We will also be able to identify genes that are controlled by EZH2 as muscle is repaired, and so may be able target these genes as therapies for improving muscle strength.