Model systems for hematologic disorders caused by ribosomal protein deficiency

描述（由申请人提供）：该申请涉及广泛的挑战领域（15）转化科学和特定的挑战主题15- dk -106：翻译基本血液学概念。核糖体蛋白（rp）基因的单倍性不足是遗传性骨髓衰竭综合征Diamond-Blackfan贫血（DBA）和5q-骨髓增生异常综合征（5q- MDS）的基础。虽然已知核糖体蛋白表达缺陷导致核糖体生物发生中断，但这种缺陷导致骨髓衰竭和MDS的分子机制尚不清楚。这种理解的缺乏限制了我们辨别当前疗法的作用机制的能力，并阻碍了这些疾病的新疗法的发展。人们普遍认为，细胞在细胞周期中前进的能力与新生核糖体的生物发生密切相关。新生核糖体生物发生的上调有两个主要功能：(i)它提供了细胞达到进入细胞周期的临界质量所需的增加的翻译能力，（ii）它确保每个子细胞获得适当数量的核糖体。鉴于核糖体与mRNA比例的变化最终会改变翻译模式和遗传程序，导致异常的生长形式，越来越多的证据表明存在检测和响应新生核糖体生物发生状态的检查点。事实上，早期的研究已经提出了p53肿瘤抑制基因的作用，最近的研究结果表明，p53在rps产生不足的情况下被激活，由于60S rpL11和rpL5结合和抑制人类双分钟2 （HDM2）而导致细胞周期阻滞。已经证明，这种检查点反应是由rpL11和rpL5通过不同的机制介导的，这取决于40S或60S rp是否被耗尽。本研究将验证的假设是，rpL11/rpL5检查点对rp应激的反应在DBA和5q- MDS的造血衰竭中起因果作用，目前的医学治疗针对这一途径以获得治疗效果。为了解决这一假设，将进行以下目标：目标1：表征rpL5和rpL11在小核糖体亚基蛋白或大核糖体亚基蛋白缺失的人造血细胞中的翻译状态；目的2：确定地塞米松、来那度胺和赖氨酸对核糖体蛋白缺失的人造血细胞中p53反应和rpL5、rpL11翻译的影响；目的3：评估地塞米松、来那度胺和亮氨酸在体内核糖体蛋白缺乏模型中的作用，并在表型患者样本中验证这些发现。从这些研究中获得的知识对于确定新的治疗靶点和开发新的合理设计的治疗方法至关重要。

DESCRIPTION (provided by applicant): This application addresses broad Challenge Area (15) Translational Science and specific Challenge Topic 15-DK-106: Translating basic hematology concepts. Haploinsufficiency of ribosomal protein (rp) genes underlies the inherited marrow failure syndromes Diamond-Blackfan anemia (DBA) and the 5q- myelodysplastic syndrome (5q- MDS). Although it is known that deficiencies in ribosomal protein expression result in disrupted ribosome biogenesis, the molecular mechanisms by which such defects lead to marrow failure and MDS are unclear. This lack of understanding limits our ability to discern the mechanism of action of current therapies and hampers the development of new therapies for these disorders. It is accepted that the ability of cells to progress through the cell cycle is tightly linked to nascent ribosome biogenesis. The upregulation of nascent ribosome biogenesis serves two major functions: (i) it provides the increased translational capacity required for a cell to achieve the critical mass to enter the cell cycle, and (ii) it ensures that each daughter cell obtains the appropriate number of ribosomes. Given that a change in the ratio of ribosomes to mRNA would eventually modify the pattern of translation and the genetic program, resulting in aberrant forms of growth, evidence is mounting that checkpoints exist to sense and respond to the status of nascent ribosome biogenesis. Indeed, earlier studies had suggested a role for the p53 tumor suppressor gene, and more recent results show that p53 is activated in response to deficiencies in the production of rps, leading to cell- cycle arrest due to the binding and inhibition of human double minute 2 (HDM2) by 60S rpL11 and rpL5. It has been demonstrated that this checkpoint response is mediated by rpL11 and rpL5 by distinct mechanisms, depending on whether a 40S or 60S rp is depleted. The hypothesis that will be tested in this study is that the rpL11/rpL5 checkpoint response to rp stress plays a causal role in hematopoietic failure in DBA and 5q- MDS and that current medical therapies target this pathway for therapeutic gain. To address this hypothesis, the following aims will be carried out: Aim 1: Characterize rpL5 and rpL11 translation status in human hematopoietic cells depleted of either small or large ribosomal subunit proteins; Aim2: Determine the effects of dexamethasone, lenalidomide, and leucine on the p53 response and on rpL5 and rpL11 translation in primary human hematopoietic cells depleted of ribosomal proteins; and Aim 3: Assess the effects of dexamethasone, lenalidomide, and leucine in an in vivo model of ribosomal protein deficiency and validate those findings in phenotyped patient samples. The knowledge derived from these studies will be critical for identifying novel therapeutic targets and for the development of new rationally designed therapies. PUBLIC HEALTH RELEVANCE: Mutations in ribosomal protein genes affect the production of ribosomes, the machinery inside the cell which is responsible for the synthesis of proteins, resulting in disorders of the hematopoietic system, such as anemia, a defect in the production of red blood cells. Although therapies are available for these diseases, their mechanism of action is not known. It is likely that the anemia of patients with mutations in genes for ribosomal proteins is due to the fact that defects in the production of ribosomes cause an arrest in cell division which can result in cell death. The aim of the proposed project is to determine the effects of therapies used to treat anemias caused by mutations in ribosomal protein genes and to identify mechanisms that trigger arrest in cell proliferation and cell death when ribosome production is impaired. This will allow for the identification of the cellular processes that impair production of red blood cell and will be essential in the design of new therapies.