Functional characterization of histone replacement variant MacroH2A1 (H2afy) in acute myeloid leukemia (AML)

组蛋白替代变体 MacroH2A1 (H2afy) 在急性髓系白血病 (AML) 中的功能特征

• 批准号: 505859092
• 负责人: Professor Dr. Florian Heidel
• 金额: --
• 依托单位: Klinik für Hämatologie, Hämostaseologie, Onkologie und Stammzelltransplantation
• 依托单位国家: 德国
• 项目类别: Research Grants
• 资助国家: 德国
• 项目状态: 未结题
• 来源: https://gepris.dfg.de/gepris/projekt/505859092?language=en
• 关键词: Functional; characterization; histone; replacement; variant

Changes of the epigenetic landscape can lead to massively altered expression patterns either directly by loss of regulatory control or through indirect, additive effects, ultimately leading to transcriptional changes. Epigenetic alterations include DNA methylation, chromatin-associated RNAs, histone tail modifications, non-histone proteins and histone variants. Among core histones, the H2A family exhibits highest sequence divergence resulting in the largest number of variants known. Amongst all H2A variants, macroH2A variants exhibit the most unique structural organization as they harbor a non-histone region at the C-terminus, named the macrodomain, making them the largest known histones. MacroH2A1 (H2afy) is highly expressed in AML patient blasts and also in hematopoietic stem cells. We have investigated the functional role of H2afy in AML and normal HSC through CRISPR-mediated inactivation and newly generated conditional mouse models. Our preliminary data provides first evidence for a functional relevance of H2afy in cell competition and maintenance of acute myeloid leukemia stem cells (AML-LSC) while it appears dispensable for normal hematopoietic stem- and progenitor cells (HSPCs). In this project we aim to determine to which extent macrohistone variant H2afy influences LSC and HSC function and how this is influenced by changes in transcription, binding partners or accessibility of specific chromatin regions. We will (1) assess for the impact of macrohistone variants on leukemic stem cell (LSC) function using MLLr-driven models of AML using rescue experiments with truncated H2afy to define relevant domains, CRISPR-mediated genome editing to define H2afy-dependent effectors and xenograft models to validate the findings in human AML. Also we aim to (2) determine the impact of genetic H2afy splicing isoforms on maintenance of normal hematopoietic stem and progenitor cells (HSPCs) and (3) analyze the mechanistic impact of macrohistone variant H2afy expression on transcription, chromatin conformation, histone modifications and binding partners in AML-LSC. Taken together, experiments described in our research proposal will produce a detailed characterization of the functional role and mechanistic impact of H2A histone variant H2afy on AML leukemia stem cells. These experiments will incorporate detailed functional analyses and genome wide characterization to provide a better understanding of biological heterogeneity induced by the presence or expression of the histone replacement variant macroH2A1 (H2afy) and prospectively explore potential therapeutic use.

表观遗传景观的变化可以通过失去调控直接或间接的加性效应导致表达模式的大规模改变，最终导致转录变化。表观遗传改变包括DNA甲基化、染色质相关rna、组蛋白尾部修饰、非组蛋白和组蛋白变体。在核心组蛋白中，H2A家族表现出最高的序列差异，导致已知变异数量最多。在所有H2A变体中，macroH2A变体表现出最独特的结构组织，因为它们在c端含有一个称为macrodomain的非组蛋白区域，使其成为已知最大的组蛋白。MacroH2A1 （H2afy）在AML患者原细胞和造血干细胞中高度表达。我们通过crispr介导的失活和新生成的条件小鼠模型研究了H2afy在AML和正常HSC中的功能作用。我们的初步数据为H2afy在急性髓系白血病干细胞（AML-LSC）的细胞竞争和维持中的功能相关性提供了第一个证据，而它在正常的造血干细胞和祖细胞（HSPCs）中似乎是必不可少的。在这个项目中，我们的目标是确定大组蛋白变体H2afy在多大程度上影响LSC和HSC的功能，以及这是如何受到转录、结合伙伴或特定染色质区域可及性变化的影响。我们将(1)使用mllr驱动的AML模型评估大组蛋白变异对白血病干细胞（LSC）功能的影响，使用截断H2afy的拯救实验来定义相关结构域，crispr介导的基因组编辑来定义H2afy依赖性效应物，以及异种移植模型来验证人类AML中的发现。此外，我们的目标是(2)确定遗传H2afy剪接异构体对正常造血干细胞和祖细胞（HSPCs）维持的影响；(3)分析大组蛋白变体H2afy表达对AML-LSC的转录、染色质构象、组蛋白修饰和结合伙伴的机制影响。综上所述，我们研究计划中描述的实验将详细描述H2A组蛋白变体H2afy对AML白血病干细胞的功能作用和机制影响。这些实验将包括详细的功能分析和全基因组表征，以更好地了解由组蛋白替代变体macroH2A1 （H2afy）的存在或表达引起的生物异质性，并前瞻性地探索潜在的治疗用途。