Capturing the dynamic epigenome using single molecule and single cell approaches

摘要 我们的身体由数十亿个基因相同的细胞组成， 不同的表型或表观遗传状态。表观遗传状态的建立，在某种程度上， 依赖于DNA包装蛋白的共价和可逆修饰， 组蛋白组蛋白H3赖氨酸9甲基化（H3K9me）与转录调控相关。 基因沉默和异染色质形成。这个过程支持正常的着丝粒 端粒功能、转座子沉默和重复DNA的失活 元素细胞中异染色质的丢失与染色体分离有关 缺陷和基因组不稳定性。我们的长期目标是识别和重建 异染色质依赖蛋白质相互作用，并捕捉这些动态 这些过程改变了单细胞和个体谱系中可遗传的基因表达状态。 虽然组蛋白在DNA复制过程中在子链之间分配， 目前尚不清楚这些修改本身是否足以作为载体 表观遗传记忆流行的观点是酶活性的平衡 能够读写和擦除组蛋白的蛋白质之间的相互作用 和表观遗传状态的遗传性。我们最近的研究表明蛋白质 以前被认为是组蛋白修饰酶的蛋白质起着结构或非结构的作用， 酶的作用，深刻影响异染色质的稳定性。利用裂变酵母， 粟酒裂殖酵母，作为一个模型系统，本建议的主要目标是1） 生物化学定义和重建异染色质依赖的蛋白质相互作用 2）确定协调父母的因素， 组蛋白转移与DNA复制，以确保表观遗传记忆传递从 在细胞分裂后，亲本细胞到子细胞3）捕获细胞轨迹 导致建立特设或适应性表观遗传状态，以应对急性 基因组应激我们希望我们的研究将确定新的调节机制， 影响异染色质组装和遗传， 利用这些途径来应对基因组内稳态的急剧变化。 .

Abstract Our bodies consist of billions of genetically identical cells with the capacity to exhibit distinct phenotypic or epigenetic states. The establishment of epigenetic states, in part, depends on the covalent and reversible modification of DNA packaging proteins called histones. Histone H3 lysine 9 methylation (H3K9me) is associated with transcriptional gene silencing and heterochromatin formation. This process underpins normal centromere and telomere function, transposon silencing and the inactivation of repetitive DNA elements. The loss of heterochromatin in cells is associated with chromosome segregation defects, and genome instability. Our long term goal is to identify and reconstitute heterochromatin dependent protein interactions and capture how these dynamic processes alter heritable gene expression states in single cells and individual lineages. Although histones are partitioned between daughter strands during DNA replication, it remains unclear as to whether the modifications themselves are sufficient to act as carriers of epigenetic memory. The prevailing notion is that a balance of enzymatic activities between proteins that can read, write and erase histone modifications alters the stability and heritability of epigenetic states. Our recent studies have revealed that proteins previously thought to act as histone modifying enzymes proteins play structural or non- enzymatic roles that profoundly impact heterochromatin stability. Using fission yeast, Schizosaccharomyces pombe, as a model system, the major goals of this proposal are 1) to biochemically define and reconstitute heterochromatin dependent protein interactions that are implicated in epigenetic inheritance 2) identify factors that coordinate parental histone transfer with DNA replication to ensure that epigenetic memory is passed on from parental cells to daughter cells following cell division 3) capture the cellular trajectory leading to the establishment of ad hoc or adaptive epigenetic states in response to acute genomic stress. We expect that our studies will identify new regulatory mechanisms that impact heterochromatin assembly and inheritance in addition to discovering how cells leverage these pathways to respond to acute changes in genomic homeostasis. .