Regulation of Ribosome Biogenesis in Hematopoietic Stem Cells

造血干细胞核糖体生物合成的调控

• 批准号: 10265594
• 负责人: Wei Tong
• 金额: $ 60.24万
• 依托单位: CHILDREN'S HOSP OF PHILADELPHIA
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-18 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10265594
• 关键词: Affect; Allogenic; Biochemical; Biogenesis; Biological; Blood; Blood Cells; Bone Marrow; Bone Marrow Transplantation; Cell Line; Cell Maintenance; Cell physiology; Cells; Clinical; Clinical Treatment; Collaborations; Cytoplasm; Data; Defect; Disease; Equilibrium; Etiology; Exhibits; Failure; Functional disorder; Gene Proteins; Genes; Genetic; Genetic Suppression; Genomics; Goals; Growth; Hematopoiesis; Hematopoietic; Hematopoietic stem cells; Human; Knockout Mice; Link; Malignant Neoplasms; Marrow; Mediating; Mutation; Myeloproliferative disease; Pancytopenia; Pathway interactions; Patients; Polyribosomes; Population; Predisposition; Prokaryotic Cells; Protein Biosynthesis; Proteins; Proteomics; Registries; Regulation; Research; Ribosomal Proteins; Ribosomes; Role; Shwachman-Diamond syndrome; Signal Transduction; Stem cell transplant; Structure; Syndrome; Testing; Therapeutic; Therapeutic Intervention; Translation Process Protein; Translations; Transplantation; Ubiquitin; Ubiquitination; Validation; Yeasts; base; bone marrow failure syndrome; cancer predisposition; cell growth; conditional knockout; epigenomics; exome sequencing; genetic analysis; in vivo; insight; interest; knock-down; novel; novel therapeutic intervention; novel therapeutics; polypeptide; prevent; programs; reconstitution; ribosome profiling; self-renewal; small hairpin RNA; stem cell function; stem cells; transcriptome; transcriptome sequencing; treatment strategy; ubiquitin-protein ligase

Abstract Tightly-regulated protein synthesis rates are critical for hematopoietic stem cell (HSC) maintenance and function. Mutations in ribosome proteins or genes that affect ribosome biogenesis cause “ribosomopathies”, a class of bone marrow failure (BMF) syndromes. As prominently illustrated by Shwachman-Diamond Syndrome (SDS), a BMF disease with progressive hematopoietic stem and progenitor cell (HSPC) failure and predisposition to myeloid malignancies, is driven by germline biallelic mutations in the assembly factors essential for the maturation of the 60S ribosome subunit. However, how ribosome assembly is regulated in HSCs remains poorly understood, as is its contribution to hematopoietic dysfunction. Importantly, other than allogeneic stem cell transplantation, therapeutic interventions that mitigate the HSPC defects in BMF do not exist. This application is based on our new studies that uncovered a novel role for the E3 ubiquitin ligase, HectD1, in regulating HSC function via ribosome biogenesis. Hectd1-deficient HSCs exhibit a striking defect in transplantation ability and self-renewal, concomitant with a reduction in global protein synthesis. The mechanism underlying HSC dysfunction upon Hectd1 deficiency is directly linked to aberrant ribosome assembly by ubiquitinating and regulating the stability of ZNF622, a critical biogenesis factor for the maturation of the 60S large ribosomal subunit in the cytoplasm. Depletion of HectD1 led to an accumulation of ZNF622 and the anti-association factor eIF6 on the 60S subunit, decreased 80S monosome to 60S ratio, consistent with a subunit joining defect associated with SDS-like diseases. Importantly, knockdown of ZNF622 in Hectd1-deficient cells restored protein synthesis and HSC reconstitution capacity. This finding represents a rare in vivo example of genetic suppression of HSC defects associated with dysfunctional ribosome biogenesis. The implications of this novel pathway to the etiology of HSC failure and clinical treatment of “ribosomopathies”, mandates detailed mechanistic understanding. Here, we propose comprehensive and in-depth analyses on the role of HectD1 and ZNF622 in ribosome biogenesis and HSCs. In aim 1, we propose to investigate the roles of HectD1 and ZNF622 in HSCs and how they interact to regulate HSC function, using a combination of complementary genetics, genomics, and biochemical approaches. In aim 2, we will systematically analyze if HectD1/ZNF622 affects different aspects of protein translation controls. Moreover, we will perform quantitative proteomics to assess if ribosome levels or ribosome composition is affected by Hectd1/ZNF622 loss. In aim 3, we will interrogate potential dysregulation of HECTD1 and ZNF622 in human BMF syndromes and explore therapeutic potential of targeting ZNF622 for the treatment of BMF with dysfunctional ribosome biogenesis. Our study implicates a previously unappreciated role of ubiquitination in regulating HSC function via controlling ribosome biogenesis factors, which are dysregulated in ribosomopathies. Our findings will likely have significant impact on the therapeutic potential of modulating ubiquitination and/or ribosome biogenesis factors in restoring HSC functions in BMF syndromes.

摘要 严格调控的蛋白质合成速率对造血干细胞(HSC)的维持和功能至关重要。 影响核糖体生物发生的核糖体蛋白或基因突变会导致核糖病，这是一类 骨髓衰竭(BMF)综合征。正如Shwachman-Diamond综合征(SDS)所突出说明的那样， 进行性造血干细胞和祖细胞(HSPC)衰竭和易感性的BMF病 髓系恶性肿瘤是由胚系双等位基因突变所致，组装因子对 60S核糖体亚基的成熟。然而，在造血干细胞中核糖体组装是如何被调控的仍然很差。 理解，以及它对造血功能障碍的贡献。重要的是，除了异基因干细胞 移植后，不存在减轻BMF中HSPC缺陷的治疗干预措施。此应用程序是 根据我们的新研究，发现了E3泛素连接酶HectD1在调节HSC中的新作用 通过核糖体生物发生发挥作用。Hectd1缺陷的HSCs在移植能力和 自我更新，伴随着全球蛋白质合成的减少。HSC的潜在机制 Hectd1缺乏症的功能障碍直接与泛素化和核糖体组装异常有关 调控ZNF622的稳定性，ZNF622是60年代大核糖体成熟的关键生物发生因子 细胞质中的亚单位。HectD1的缺失导致ZNF622和抗结合因子的积累 60S亚基上的eIF6降低了80s单体与60s的比率，与亚基连接缺陷一致 与类抑郁疾病有关。重要的是，在Hectd1缺陷细胞中敲除ZNF622恢复了蛋白质 合成和HSC重建能力。这一发现代表了一个罕见的体内基因抑制的例子。 与核糖体生物发生功能障碍相关的HSC缺陷。这一新途径的含义是 HSC衰竭的病因和“核糖体疾病”的临床治疗，要求详细的机制 理解。在这里，我们提出了全面和深入的分析HectD1和ZNF622在 核糖体生物发生与肝干细胞。在目标1中，我们建议研究HectD1和ZNF622在HSCs中的作用 以及它们如何相互作用来调节HSC功能，使用互补遗传学、基因组学和 生化方法。在目标2中，我们将系统分析HectD1/ZNF622是否会影响到 蛋白质翻译控制。此外，我们将进行定量蛋白质组学来评估核糖体水平或 核糖体组成受Hectd1/ZNF622缺失的影响。在目标3中，我们将询问潜在的失调 HECTD1和ZNF622在人BMF综合征中的表达，并探讨靶向ZNF622治疗BMF综合征的可能性 核糖体生物发生功能障碍治疗BMF我们的研究暗示了一个以前未被认识到的角色 泛素化通过调控核糖体生物发生因子调控肝星状细胞功能 在核糖体疾病中。我们的发现可能会对调节药物的治疗潜力产生重大影响。 泛素化和/或核糖体生物发生因子在BMF综合征恢复HSC功能中的作用