Histone isomerization and pluripotency

描述（由申请人提供）： 成功的体细胞核克隆已经证明，卵细胞质中含有强大的核重编程活性。这些活性可以诱导终末分化的供体细胞核的去分化，导致它们获得全能状态。尽管克隆研究的历史悠久，特别是最近的研究密集，然而，很少有人知道在克隆过程中重新编程供体细胞核的分子机制。该项目的长期目标是鉴定这些卵源性核重编程因子，并应用这些因子重编程活哺乳动物细胞的分化状态。该项目有望为自体移植有效制备患者来源的新组织，补充诱导多能干细胞（iPS）技术。 整体染色质去凝聚是在供体细胞核中观察到的最显著的细胞生物学变化之一。研究小组最近发现，卵蛋白质核质溶酶和肽基异构酶p16负责染色质解凝聚。这两种蛋白质触发特定组蛋白上的乙酰化、磷酸化和甲基化，并促进基因活化。p16和核质蛋白的转导显著促进小鼠成纤维细胞产生iPS细胞。基于这些观察，假设核质蛋白和p16在靶基因上组织了一组特定的表观遗传修饰，这导致染色质松弛和核重编程。为了验证这一假设，提出了以下具体目标。具体目标1旨在了解核浆蛋白和p16如何在iPS细胞形成期间促进关键多能性基因的表观遗传重编程。在Specific Aim 2中，将通过基因敲除、DNA微阵列分析和染色质免疫沉淀来研究p16在ES细胞多能性中的作用。在具体目标3中，将通过使用肽片段、染色质和活细胞来研究组蛋白的磷酸化、甲基化和异构化之间的关系。总之，这些研究有望为通过表观遗传重塑重新编程细胞分化提供新的见解。 公共卫生相关性： 这些研究将促进对干细胞生物学中染色质修饰和细胞重编程的机制理解。这对于高效的 以及干细胞在再生医学中的安全临床应用。

DESCRIPTION (provided by applicant): Project Summary Successful somatic cell nuclear cloning has demonstrated that egg cytoplasm contains powerful nuclear reprogramming activities. These activities can induce dedifferentiation of the terminally differentiated donor nuclei, leading to their acquisition of a totipotent state. Despite the long history of the cloning study and particularly intensive recent research, however, little is known about the molecular mechanisms that reprogram donor nuclei during cloning. The long-term goal of this project is to identify these egg-derived nuclear reprogramming factors and apply the factors to reprogram the differentiation status of living mammalian cells. This project i expected to lead to efficient preparation of patients-derived new tissues for autologous transplantation, complementing the induced pluripotent stem (iPS) cell technology. Global chromatin decondensation is one of the most striking cell biological changes observed in the donor nuclei. The investigator's group has recently found that the egg protein nucleoplasmin and the peptidyl isomerase p16 are responsible for the chromatin decondensation. These two proteins trigger acetylation, phosphorylation and methylation on specific histones and promote gene activation. Transduction of p16 and nucleoplasmin significantly facilitate the production of iPS cells from mouse fibroblasts. Based on these observations, it was hypothesized that nucleoplasmin and p16 organize a specific set of epigenetic modifications on the target genes, which leads to chromatin relaxation and nuclear reprogramming. To test this hypothesis, following specific aims are proposed. Specific Aim 1 is designed to understand how nucleoplasmin and p16 contributes to epigenetic reprogramming of key pluripotency genes during iPS cell formation. In Specific Aim 2, the role of p16 in pluripotency of ES cells will be studied by gene knockdown, DNA microarray analysis and chromatin immunoprecipitation. In Specific Aim 3, the relationship among phosphorylation, methylation and isomerization of histones will be investigated by using peptide fragments, chromatin and living cells. Together, these studies are expected to provide a novel insight into the reprogramming of cell differentiation through epigenetic remodeling. PUBLIC HEALTH RELEVANCE: Project Narrative These studies will facilitate mechanistic understanding of chromatin modifications and cellular reprogramming in stem cell biology. This is also important for efficient and safe clinical applications of stem cells in regenerative medicine.