Function of RUNX1 in diverse Down syndrome tissues

项目概要 家长提案：RUNX1，也称为急性髓系白血病 1 蛋白 (AML1)，是一种 在造血谱系规范中发挥关键作用的转录因子 在胚胎发生期间，是整个出生后正常巨核细胞生成所必需的 生活。 RUNX1 基因定位于 21 号染色体的带 q22.12，在 患有 21 三体症 (T21) 的个体。因此，提出了 RUNX1 的额外副本 在与唐氏综合症（DS）相关的血液学改变中发挥相关作用。我们 试图破译 RUNX1 对唐氏综合症患者表型的贡献 综合症。由于转录因子与 DNA 结合并改变 RNA 聚合酶活性，我们将 确定 RUNX1 的这两个功能在唐氏综合症衍生细胞中是否发生改变。我们会 还定义了增加或抑制 RUNX1 功能的药物在具有 RUNX1 功能的细胞中是否表现不同 21号染色体的额外拷贝。最后，我们将确定有多少被改变的血液 唐氏综合症中所见的分化是由 RUNX1 通过“正常化”RUNX1 基因引起的 三体背景中的剂量（例如，三体细胞系中的两个拷贝）。然后我们将分析 iPSC 分化为胚状体和血细胞。总的来说，这项工作将摆脱 对 RUNX1 功能及其在 21 三体中如何改变的重要见解。 补充项目摘要：Gialesaki 等人最近的一份报告。 （2023年3月）成立 RUNX1 选择性剪接和异构体不平衡的关键作用 - 而不是整体 表达-作为唐氏综合症相关骨髓性白血病（DS-ML）的主要驱动因素。这些 研究结果对我们提案的大多数具体目标具有重要影响。因此，我们 应该部分修改我们的一些子目标来评估 RUNX1 剪接的相关性 DS 来源的 iPSC 和类淋巴母细胞分化的不同结果的平衡 和体内平衡。此补充的第二个动机仅适用于目标 3。此处 我们最初提出通过敲除 T21 中的一个拷贝来标准化 RUNX1 基因剂量 / /− iPSC 线 (T21 RUNX1)。我们现在提出替代策略来克服这种问题 障碍并结合刚才提到的异构体特异性分析。本行政 补充内容涉及第 1 部分：有针对性的高风险高回报基础科学研究 这与唐氏综合症的白血病风险相关。此外，这种补充将显着 通过促进更好地理解该项目来提高该项目的科学意义 控制 T21 个体血液学改变的潜在机制。

Project Summary Parent Proposal: RUNX1, also known as Acute Myeloid Leukemia 1 protein (AML1), is a transcription factor that plays a critical function in the specification of the hematopoietic lineage during embryogenesis and is required for normal megakaryopoiesis throughout the postnatal life. The RUNX1 gene is localized to band q22.12 of chromosome 21, which is triplicated in individuals with Trisomy 21 (T21). For this reason, an extra copy of RUNX1 has been proposed to play relevant roles in the hematological alterations associated with Down syndrome (DS). We seek to decipher the contribution of RUNX1 to the phenotypes seen in individuals with Down syndrome. Since transcription factors bind to DNA and alter RNA polymerase activity, we will determine if those two functions of RUNX1 are altered in Down syndrome-derived cells. We will also define if drugs that increase or inhibit RUNX1 function behave differently in cells with an extra copy of chromosome 21. Finally, we will ascertain how much of the altered blood differentiation seen in Down syndrome is caused by RUNX1 by ‘normalizing’ RUNX1 gene dosage in a trisomy background (e.g., two copies in a trisomy cell line). We will then analyze the differentiation of iPSCs into embryoid bodies and blood cells. Collectively this work will shed important insights into the functions of RUNX1 and how it is altered in Trisomy 21. Supplement Project Summary: A recent report by Gialesaki et al. (March 2023) established a critical role for RUNX1 alternative splicing and isoform imbalance -rather than overall expression- as a major driver of Down syndrome-associated myeloid leukemia (DS-ML). These findings have important implications on most of the specific aims of our proposal. Therefore, we should partially modify some of our sub-aims to assess the relevance of RUNX1 splicing equilibrium on the different outcomes of DS-derived iPSC and lymphoblastoid cell differentiation and homeostasis. A second motivation for this supplement applies exclusively to Aim 3. Here we orginally proposed to normalize RUNX1 gene dosage by knocking-out one copy in the T21 +/+/− iPSC line (T21 RUNX1 ). We now propose alternative strategies to overcome such obstacles and incorporate the isoform-specific analyses just mentioned. This Administrative Supplement addresses Component 1: A targeted high risk - high reward basic science study that is relevant to Leukemia risk in Down syndrome. Moreover this supplement will significantly improve the scientific significance of the project by contributing to a better understanding of the underlying mechanisms that govern hematological alterations in individuals with T21.