The role of Kmt2c/MLL3 in hematopoietic stem cell self-renewal, commitment and exhaustion

项目摘要 本提案的目标是了解KMT 2C/MLL 3如何调节造血干细胞（HSC）的自我调节。 更新和为什么KMT 2C突变传达了可导致白血病的选择性优势。KMT 2C编码 MLL 3，COMPASS家族组蛋白甲基转移酶，结合增强子元件并促进转录。 KMT 2C在人类白血病中突变，既作为染色体7 q大缺失的一部分，也作为特异性染色体缺失的一部分。 结构域如PHD锌指结构域（其结合染色质）或SET甲基转移酶结构域 （其可以引发增强子元件以用于激活）。先前的小鼠研究已经确定Kmt 2c缺失 增强HSC自我更新，促进白血病发生，但其机制尚不清楚。更好地了解 为了研究Kmt 2c如何调节HSC自我更新，我们产生了种系和条件性功能丧失小鼠。Kmt2c 缺失增强了HSC的自我更新，与先前的观察结果一致，但突变并没有改变细胞的功能。 自行循环动力学。相反，Kmt 2c缺失允许连续移植或化疗治疗， HSC在多次分裂循环后保持自我更新能力。这使得突变的HSC在竞争中胜出 在骨髓恢复期间的野生型HSC。在没有压力的情况下，Kmt 2c缺失并不表达选择性的表达。 优势总之，我们的数据表明，Kmt 2c突变减轻了一种称为HSC耗竭的现象， 其中HSC在几次累积分裂后失去自我更新能力。我们的机械数据表明， MLL 3通过以下任一途径引发HSC响应于IL-1和可能的其他炎性细胞因子而分化： 增强IL-1信号转导或促进IL-1靶基因表达。这项建议的目的是 旨在扩展这些观察。目的1将测试Kmt 2c/MLL 3缺陷是否传递选择性的免疫应答。 通过降低对IL-1和其他炎性细胞因子的敏感性，这有利于分裂HSC。目标2将 表征HSC中MLL 3靶增强子的结构、调节和IL-1反应性， 有着丰富的组织历史增强子引发的变化可以允许HSC存档它们的分裂历史， 在多个分裂周期后，更倾向于承诺，而不是自我更新。目标3将测试MLL 3是否需要 功能性SET或PHD结构域以限制HSC自我更新能力。这种结构-功能分析将有助于 我们更好地了解特定的KMT 2C突变如何传递自我更新优势。如果我们能 了解HSC在经历累积的自我更新分裂时如何变化，以及Kmt 2c缺失如何发生 传达选择性自我更新的优势，我们最终可能能够通过时期来保留HSC功能。 压力，如化疗后时期，而不会增加白血病的风险。

PROJECT SUMMARY The goal of this proposal is to understand how KMT2C/MLL3 regulates hematopoietic stem cell (HSC) self- renewal and why KMT2C mutations convey a selective advantage that can lead to leukemia. KMT2C encodes MLL3, a COMPASS family histone methyltransferase that binds enhancer elements and promotes transcription. KMT2C is mutated in human leukemias, both as part of large deletions of chromosome 7q and at specific domains such as the PHD zinc finger domains (which bind chromatin) or the SET methyltransferase domain (which can prime enhancer elements for activation). Prior murine studies have established that Kmt2c deletions enhance HSC self-renewal and promote leukemogenesis, but the mechanism is not clear. To better understand how Kmt2c regulates HSC self-renewal, we generated germline and conditional loss-of-function mice. Kmt2c deletions enhanced HSC self-renewal, consistent with prior observations, but the mutations did not alter cell cycle kinetics by themselves. Instead, Kmt2c deletions allowed serially transplanted or chemotherapy treated HSCs to retain self-renewal capacity after multiple division cycles. This allowed the mutant HSCs to outcompete wild type HSCs during marrow recovery. In the absence of stress, Kmt2c deletions did not convey a selective advantage. Altogether, our data suggest that Kmt2c mutations mitigate a phenomenon, called HSC exhaustion, in which HSCs lose self-renewal capacity after several cumulative divisions. Our mechanistic data suggest that MLL3 primes HSCs to differentiate in response to IL-1, and possibly other inflammatory cytokines, by either enhancing IL-1 signal transduction or by facilitating IL-1 target gene expression. The aims of this proposal are designed to extend these observations. Aim 1 will test whether Kmt2c/MLL3 deficiency conveys a selective advantage to dividing HSCs by reducing sensitivity to IL-1 and other inflammatory cytokines. Aim 2 will characterize the structure, regulation and IL-1 responsiveness of MLL3 target enhancers in HSCs with short and extensive division histories. Changes in enhancer priming may allow HSCs to archive their division histories and favor commitment, rather than self-renewal, after multiple division cycles. Aim 3 will test whether MLL3 requires functional SET or PHD domains to restrict HSC self-renewal capacity. This structure-function analysis will help us better understand how specific KMT2C mutations might convey a self-renewal advantage. If we can understand how HSCs change as they undergo cumulative self-renewing divisions, and how Kmt2c deletions convey a selective self-renewal advantage, we may ultimately be able to preserve HSC function through periods of stress, such as post-chemotherapy periods, without increasing leukemia risk.