Deciphering the regulatory principles governing enhancer specificity

摘要 增强子是基因组元件，其编码基因何时和在何处表达的指令。 在发展过程中表现出来。大多数导致疾病的突变被认为存在于 增强剂。然而，我们并不了解增强子序列中哪些变化是惰性序列 个体或群体之间的差异，影响基因调控和细胞完整性。 这些基本问题仍然没有解决，因为我们不能将增强子序列与基因 表达和表型。我们知识上的这一差距阻碍了我们解释基因组数据的能力 了解发育、细胞完整性和疾病。增强子序列提供了一个支架， 通过识别DNA中的特定签名来结合转录因子。的物理约束 控制这些蛋白质如何与增强子DNA相互作用的基因， 可用于理解增强子序列和组织之间的关系的约束 特异性基因表达我提议开发一套方法和办法， 破译组织特异性增强子活性的语法限制。我将使用高度并行 功能性报告基因分析、高通量基因型-表型研究沿着生物化学 分析、合成生物学和功能丧失策略。这些实验将在 脊索玻璃海鞘，因为它是一个独特的系统，其中数百万个增强子变体可以被 对发育中的胚胎的所有细胞的功能进行分析。在Ciona的工作将得到补充， 在鸡、小鼠和组织培养中进行实验，直接告知脊椎动物的发育情况， 突变导致疾病。确定与基因编码序列相关的“遗传密码” 对蛋白质的研究提供了对我们基因组主要组成部分的详细了解。一旦我们有了 类似的代码来破译这些基因何时何地表达的指令，我们将有 强大的工具来了解基因组如何编码构建和维护 生活

Abstract Enhancers are the genomic elements that encode the instructions for when and where genes are expressed during development. The majority of mutations leading to disease are thought to reside in enhancers. However, we do not understand which changes in enhancer sequence are inert sequence variations between individuals or populations and which impact gene regulation and cellular integrity. These fundamental questions remain unsolved, because we cannot relate enhancer sequence to gene expression and phenotype. This gap in our knowledge is stalling our ability to interpret genomic data and understand development, cellular integrity, and diseases. Enhancer sequences provide a scaffold for transcription factors to bind to, by recognizing specific signatures in the DNA. The physical constraints that govern how these proteins interact with enhancer DNA could lead to a set of grammatical constraints that can be used to understand the relationship between enhancer sequence and tissue specific gene expression. I propose the development of a toolkit of methodologies and approaches to decipher the grammatical constraints on tissue specific enhancer activity. I will use highly parallel functional reporter assays, high-throughput genotype to phenotype studies along with biochemical assays, synthetic biology, and loss of function strategies. These experiments will be carried out in the chordate Ciona intestinalis, as it is a unique system in which millions of enhancer variants can be assayed for function in all cells of a developing embryo. Work in Ciona will be complemented with experiments in chick, mouse and tissue culture to directly inform vertebrate development and pinpoint mutations causing disease. Determining the ‘genetic code’ that relates the coding sequences of genes into protein has provided detailed insight into a major component of our genome. Once we have a similar code to decipher the instructions for when and where these genes are expressed, we will have powerful tools to understand how the genome encodes the instructions for building and maintaining life.