Understanding how enhancer chromatin transduces extracellular signalling during developmental transitions in human pluripotent cells

一个细胞的身份是由它的基因决定的。例如，位于心脏的细胞将表达心脏基因，但不表达对肝脏或大脑功能重要的基因。相反，肝细胞表达肝脏基因，但不表达心脏或肺基因。在胚胎发育期间，动物由不同的干细胞群形成，这些干细胞能够产生在成年生物体中发现的无数不同的细胞类型。细胞“知道”它们应该成为哪种细胞类型，因为它们接收到的信号通常是由相邻细胞发送的，以便在正确的时间处于正确位置的细胞具有适当的身份。当这些信号的水平达到某些阈值时，做出适当的反应不仅在发育过程中而且在维持健康状态方面都至关重要。当这些反应失败时，后果包括发育障碍和癌症。当干细胞经历分化成新细胞类型的过程时，它们需要打开适合新细胞类型的基因，但也需要关闭定义“干细胞”身份的基因，并阻止任何与其他细胞类型相关的基因。这一过程受到一组称为染色质重塑蛋白质的严格调控。DNA在细胞内并不是“裸露的”，而是以染色质的形式存在，即它以一种保持紧凑和稳定的方式包裹在蛋白质周围。染色质重塑剂能够改变染色质的特定调节部分的紧密程度。因此，如果细胞接收到正确的信号，染色质重塑因子就能使一些基因做出反应，而另一些基因则保持沉默。我们感兴趣的是，进入细胞的信号如何被解释为基因表达的变化。我们最近在小鼠多能细胞中发现了一个以前未被检测到的步骤，该步骤原本是一个研究得很好的信号通路。这些细胞中的信号激活导致非常快的事件，其中关键调控序列的染色质从"等待“构型重置为”活跃“构型。我们进一步表明，不同的染色质重塑蛋白质所需的不同步骤，在这个重置事件。我们现在希望确定人类多能细胞如何响应这些相同信号的激活。我们目前的数据表明，重置事件发生在人类细胞中，但我们已经检测到物种特异性差异。在目前的研究中，我们将超越小鼠工作的范围，以确定细胞如何响应不同信号通路水平的变化，并确定为什么有些细胞对信号有反应，而另一些则没有。我们将利用我们所学到的知识来设计控制细胞对信号的反应性的方法，这将在再生医学和治疗细胞产生异常信号反应的疾病（如癌症）方面带来巨大的好处。我们正在使用尖端的方法分析染色质行为在全基因组水平和单分子水平。总的来说，这项提议代表了对早期胚胎人类细胞如何定向形成成年生物体中存在的无数细胞类型的全面调查，通过这些调查，我们将设计出控制这一过程的方法。

A cell's identity is defined by its genes. A cell located in the heart, for example, will express heart genes, but not genes important for liver or brain function. Conversely, liver cells express liver genes but not heart or lung genes. During embryonic development animals are formed by different groups of stem cells, which are able to give rise to the myriad of different cell types found in an adult organism. Cells "know" which cell types they should become because of signals they receive, usually sent by neighbouring cells so that cells in the right place at the right time take on the appropriate identity. Responding appropriately when the levels of these signals reach certain thresholds is crucial not only during development but also in maintaining a healthy state. When these responses fail the consequences include developmental disorders and cancer.When stem cells undergo the process of differentiating into a new cell type, they need to turn on genes appropriate for the new cell type, but also turn off the genes that define a 'stem cell' identity and also keep off any genes associated with other cell types. This process is tightly regulated by a group of proteins called chromatin remodellers. DNA is not "naked" within cells but exists in the form of chromatin, i.e. it is wrapped around proteins in a way that keeps it compact and stable. Chromatin remodellers are able to change how tightly packed specific regulatory parts of the chromatin are. In this way chromatin remodellers are important for making some genes available to respond if the right signal is received by a cell, while other genes are kept silent.We are interested in how the signals which enter a cell are interpreted to produce a change in gene expression. We recently discovered a previously undetected step in an otherwise well studied signalling pathway in mouse pluripotent cells. Signal activation in these cells results in a very fast event in which the chromatin of key regulatory sequences gets reset from a 'waiting' configuration to an 'active' configuration. We further showed that different chromatin remodelling proteins are required for different steps in this resetting event. We now wish to determine how human pluripotent cells respond to activation of these same signals. Our current data indicates that a resetting event occurs in human cells, but we've already detected species-specific differences. In the current study we will move beyond the scope of the mouse work to define how cells respond to changes levels of different signalling pathways, and to determine why some cells are responsive to signals while others are not. We will use what we learn to devise methods for controlling the responsiveness of cells to signals, something that would be of enormous benefit in regenerative medicine and in treatment of diseases where cells exert aberrant signalling responses, such as cancer. We are using cutting-edge methodologies analysing chromatin behaviour at the genome-wide level and at the single molecule level. Overall, this proposal represents a comprehensive investigation of how early embryonic human cells are directed to form the myriad of cell types present in an adult organism, through which we will devise ways in which to control this process.