Investigating epigenetic inheritance patterns and mechanisms during asymmetric division of Drosophila female germline stem cells

表观遗传修饰提供了包含相同基因组的细胞能够 采用并维护不同的小区身份。这些机制对于维持干细胞库至关重要。 它可以分化和补充在动态平衡、损伤和衰老过程中丢失的组织。为了维持干细胞 池，不对称细胞分裂(ACD)发生在一个子细胞成为自我更新的干细胞和 另一个女儿继续区分。破坏这种平衡可能是灾难性的，会导致缺陷。 从癌症到退化性疾病。尽管它很重要，但人们对它的内容和方式知之甚少 在ACD期间，干细胞的表观遗传信息可以在其两个子体之间分布。该项目 这里提出的将(1)允许可视化包含在基因特定基因座上的表观遗传信息 分离ACD中的姐妹染色单体和(2)描述信徒中涉及的顺式和反式元件 在两个子细胞之间分离表观遗传上不同的染色单体。 表观遗传控制中的一个主要参与者是核小体，它是组蛋白的八聚体，它密切地 与DNA相互作用，并具有可广泛修饰以影响基因表达的N-末端“尾巴”。它是 众所周知，组蛋白的年龄与其携带的表观遗传修饰的类型有关。因此，它可能是 ACD的子代可能通过继承“旧的”和“新的”组蛋白而获得不同的命运 对维持干细胞状态或促进分化很重要的基因。果蝇种系 允许在体内以单细胞分辨率显示不对称生殖系干细胞(GSC)分裂，使其 这是一个研究ACD子代间组蛋白分离的伟大系统。实验室的最新数据显示 事实上，在有丝分裂的GSC中发现了新旧组蛋白信号不重叠的重要区域。去调查 在这些区域，一种新的成像技术已经被开发出来。简而言之，以基因座为靶点的荧光探针 含有‘茎’或分化基因的基因被用来确定特定的基因是否优先 与旧的和新的组蛋白相关联。此外，这些探针将与特定的抗体一起使用 组蛋白修饰以揭示更新的干细胞子代和 女儿注定要在ACD中脱颖而出。实时成像和固定成像将阐明这两个顺式元件的动力学。 关于有丝分裂机制上的染色单体和反式调节因子在姐妹染色单体中的作用 GSC ACD过程中的染色单体识别和分离。 结果将揭示基因特异性表观遗传控制是如何在ACD期间建立的，以及提供 一张分子结构和细胞事件的时空图，这些分子机械和细胞事件涉及忠实地分离 信息。这些方法可以广泛应用于其他生物和细胞系，这将增强 研究ACD的关键过程的能力。这里生成的数据和技术将显著影响 干细胞和染色质生物学领域的基础知识。

Epigenetic modifications provide essential mechanisms by which cells containing identical genomes can adopt and maintain different cell identities. These mechanisms are crucial for maintaining pools of stem cells that can differentiate and replenish tissue lost during homeostasis, injury, and aging. To maintain the stem cell pool, asymmetric cell division (ACD) occurs where one daughter cell becomes a self-renewed stem cell and the other daughter goes on to differentiate. Disruption of this balance can be disastrous, leading to defects ranging from cancer to degenerative diseases. Despite its importance, little is known about which and how the epigenetic information of the stem cell could be distributed between its two daughters during ACD. The project proposed here will (1) permit visualization of the epigenetic information contained at gene-specific loci on segregating sister chromatids during ACD and (2) describe the cis and trans elements involved in the faithful segregation of the epigenetically distinct chromatids between the two daughter cells. A major player in epigenetic control is the nucleosome, an octamer of histone proteins that intimately interacts with DNA and has N-terminal `tails' that can be extensively modified to affect gene expression. It is known that the age of a histone is correlated with the types of epigenetic modifications it carries. Thus, it may be possible that the daughters of an ACD may acquire their different fates by inheriting `old' versus `new' histones at genes important for maintaining the stem cell state or promoting differentiation. The Drosophila germline permits visualization of asymmetric germline stem cell (GSC) division at single-cell resolution in vivo, making it a great system to study histone segregation between the daughters of an ACD. Current data in the lab has indeed found significant regions of non-overlapping old and new histone signals in mitotic GSCs. To investigate these regions, a new imaging technique has been developed. In short, fluorescent probes targeting loci containing either `stemness' or differentiation genes are applied to determine if a particular gene is preferentially associated with old versus new histones. Further, these probes will be used with antibodies against specific histone modifications to reveal the epigenetic differences between the renewed stem cell daughter and the daughter destined to differentiate in ACD. Live and fixed imaging will elucidate the dynamics of both cis elements on the chromatids and trans regulatory factors on the mitotic machinery to determine the roles they play in sister chromatid recognition and segregation during GSC ACD. The results will reveal how gene-specific epigenetic control is established during ACD as well as provide a spatiotemporal map of the molecular machinery and cellular events involved in faithfully segregating that information. These methods can be widely applied to other organisms and cell lineages, which will enhance the capacity to study the crucial process of ACD. The data and techniques generated here will significantly impact fundamental knowledge in the fields of stem cell and chromatin biology.