Molecular feedback between cell division cycle and differentiation in pluripotent stem cells

项目摘要 我实验室的一个主要长期目标是确定分子反馈机制 负责协调细胞分裂周期和胚胎干细胞分化 细胞（ESC）。胚胎干细胞在再生医学中具有巨大的治疗潜力，因为它们可以 它可以分化成任何细胞类型，并具有无限的自我更新潜力。这个非凡 生物潜能，称为多能性，与复杂的转录网络有关， 一个超快的细胞分裂周期，缺乏典型的检查点，以及对激活的非典型反应 丝裂原活化蛋白激酶（MAPK）通路。虽然已经取得了很大的进展 在描述多能性转录网络时，对细胞分裂周期的了解较少 及其与多能性转录网络的分子联系。此外，一个关键的 一个知之甚少的过程是胚胎干细胞如何使用MAPK途径来控制它们的自我更新， 分化我们的假设是细胞分裂周期和多能性 转录网络通过双向分子反馈环连接， 由MAPK活性调节。为了验证我们的假设，我们将：表征新型稳压器 在我们的胚胎干细胞II中的CRISPR筛选中鉴定的MAPK通路。确定功能底物 使用化学遗传激酶工程的MAPK激酶Mek和Erk在ESC中的网络 和定量磷酸化蛋白质组学。三.确定多能性维持的机制， G1期细胞周期激酶（Cdk2），使用化学遗传学直接标记底物， Cdk2的设计为了成功完成实验，我建立了 与Stanley Qi教授实验室（斯坦福大学）、Alice Ting教授合作 实验室（斯坦福大学），Seth Rubin教授实验室（加州大学，圣 Cruz）和Boris Macek教授实验室（图宾根大学）。通过获得详细的洞察力 细胞分裂周期和胚胎干细胞分化的分子机制， 这项R35建议的结果将为解决该领域的一个关键挑战提供新的战略 再生医学是有效和可再生的胚胎干细胞分化，用于治疗 目的

Project Summary A major long-term goal of my laboratory is to determine molecular feedback mechanisms responsible for coordination between the cell division cycle and differentiation of embryonic stem cells (ESCs). ESCs have great therapeutic potential for regenerative medicine, because they can differentiate into any cell type and have unlimited self-renewal potential. This remarkable biological potential, known as pluripotency, is associated with a complex transcription network, an ultrafast cell division cycle that lacks typical checkpoints, and an atypical response to activation of the Mitogen-Activated Protein Kinase (MAPK) pathway. While a lot of progress has been made in characterizing the pluripotency transcription network, less is known about the cell division cycle of ESCs and its molecular links to the pluripotency transcription network. In addition, a critical yet poorly understood process is how ESCs use the MAPK pathway to control their self-renewal and differentiation. Our driving hypothesis is that the cell division cycle and pluripotency transcription network are linked through a bidirectional molecular feedback loop that is regulated by the MAPK activity. To test our hypothesis, we will: I. Characterize novel regulators of the MAPK pathway identified in our CRISPR screen in ESCs II. Determine functional substrate network of the MAPK kinases, Mek and Erk, in ESCs using chemical-genetic kinase engineering and quantitative phosphoproteomics. III. Determine mechanisms of pluripotency maintenance by the G1 cell cycle kinase (Cdk2) using direct labeling of the substrates by chemical-genetic engineering of Cdk2. To successfully complete proposed experiments, I have established collaborations with Professor Stanley Qi laboratory (Stanford University), Professor Alice Ting laboratory (Stanford University), Professor Seth Rubin laboratory (University of California, Santa Cruz) and Professor Boris Macek laboratory (University of Tubingen). By gaining detailed insight into the molecular mechanisms linking the cell division cycle and differentiation of ESCs, the outcome of this R35 proposal will provide novel strategies to address a key challenge in the field of regenerative medicine that is efficient and reproducible differentiation of ESCs for therapeutic purposes.