Chromatin maintenance and sister chromatid differentiation

描述（由申请方提供）：在亲本DNA模板链复制后，预期姐妹染色单体具有完全相同的DNA序列，但DNA复制本身导致的突变除外。在细胞分裂过程中，表观遗传标记如何传递给下一代细胞尚不清楚。在有丝分裂之前未丢失的表观遗传标记随机分布在姐妹染色单体之间或分配给特定的姐妹染色单体，例如取决于亲本DNA模板链是否通过前导链或滞后链DNA复制而复制。或者，姐妹染色单体可以以链特异性的方式印记。姐妹染色单体之间的染色质差异可能在功能上不重要，或者导致子细胞之间基因表达的“随机”差异。姐妹染色单体之间的表观遗传差异也可能参与细胞命运的调节，如果微管起源于“母亲”的中心体是更喜欢着丝粒目前只有一个姐妹染色单体的特定染色体。直到最近，还不可能可靠地鉴定和区分姐妹染色单体。然而，我们最近发现DNA模板链序列可以可靠地用于鉴定姐妹染色单体。这些新技术将被用来研究姐妹染色单体之间表观遗传差异的可能性及其功能意义。这些研究有三个具体目标：1。研究单细胞中遗传的特定姐妹染色单体的基因表达。2.研究含有“沃森”或“克里克”DNA模板链序列的分选姐妹染色单体上的染色质标记。3.在体外和体内跟踪姐妹染色单体从特定染色体分离成特定细胞类型。针对这些互补的具体目标的拟议研究将澄清姐妹染色单体之间是否存在染色质差异，如果存在，这种差异是否与子细胞之间基因表达的随机变异和细胞命运决定以及多细胞生物体的发育有关。重要的是，拟议的研究将回答一个以前没有解决的问题：一个给定的子细胞继承哪个姐妹染色单体重要吗？如果答案是肯定的，所提出的研究和实验方法将为进一步研究一个全新的研究领域铺平道路：染色质复制和姐妹染色单体分化。这类研究的意义很难预测，但范围从识别新的治疗靶点和开发从（诱导）多能干细胞开始的更有效的细胞疗法到靶向正常和恶性干细胞自我更新的更有效策略。

DESCRIPTION (provided by applicant): Following replication of parental DNA template strands, sister chromatids are expected to have exactly the same DNA sequence except for mutations resulting from DNA replication itself. How epigenetic marks are transmitted to the next cell generation during cell division is less clear. Epigenetic marks that are not lost prior to mitosis are either distributed randomly between sister chromatids or assigned to a specific sister chromatid e.g. depending on whether the parental DNA template strand was replicated by either leading or the lagging strand DNA replication. Alternatively, sister chromatids could be imprinted in a strand-specific manner. Chromatin differences between sister chromatids could be functionally insignificant or lead to "stochastic" differences in the expression of genes between daughter cells. Epigenetic differences between sister chromatids could also be involved in cell fate regulation if microtubules originating from "mother' centrosomes were to prefer kinetochores present on just one of the two sister chromatids of specific chromosomes. Until recently it was not possible to reliably identify and distinguish sister chromatids. However, we recently found that DNA template strand sequences can be reliably used to identify sister chromatids. These novel techniques will be used to study the possibility of epigenetic differences between sister chromatids and their functional implications. The proposed studies have three specific aims: 1. Study gene expression in relation to specific sister chromatids that were inherited in single cells. 2. Study chromatin marks on sorted sister chromatids containing either "Watson" or "Crick" DNA template strand sequences. 3. Follow the segregation of sister chromatids from specific chromosomes into specific cell types in vitro and in vivo. The proposed studies towards these complementary specific aims will clarify whether chromatin differences between sister chromatids exist and if so, whether such differences are relevant to stochastic variation in gene expression between daughter cells and cell fate decisions and the development in multicellular organisms. Importantly, the proposed studies will answer a question which has not been addressed before: does it matter which sister chromatids are inherited by a given daughter cell? If the answer is affirmative, the proposed studies and experimental approaches will pave the way for further studies of an entirely novel field of research: chromatin replication and sister chromatid differentiation. The implications of this type of research are hard to predict but range from the identification of novel targets for therapy and the development of more effective cell therapies starting from (induced) pluripotent stem cells to more effective strategies that target self-renewal in normal and malignant stem cells.