The Role of PKR in a Novel IL-3 Signal Transduction Pathway

PKR 在新型 IL-3 信号转导途径中的作用

• 批准号: 7819168
• 负责人: William Stratford MAY
• 金额: $ 1.7万
• 依托单位: UNIVERSITY OF FLORIDA
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-06-08 至 2010-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7819168
• 关键词: Acute leukemia; Apoptosis; Biochemical; Bone Marrow; Bone Marrow Cells; Cell Culture Techniques; Cell Cycle; Cell Cycle Arrest; Cells; Cellular Stress; Cellular Stress Response; Characteristics; Clinical; Data; Differentiation and Growth; Disease; Dominant-Negative Mutation; Double-Stranded RNA; Dysmyelopoietic Syndromes; Evolution; Excision; Fanconi' s Anemia; Funding; Gene Expression; Genetic Transcription; Growth Factor; Hematopoietic; Hematopoietic Cell Growth Factors; Human; Infection; Inflammatory; Inhibition of Apoptosis; Interleukin-3; Knowledge; Laboratories; Leukemic Cell; Mediating; Mitochondria; Modeling; Molecular; NUP98 gene; Outcome; Pathway interactions; Patients; Phosphorylation; Protein Biosynthesis; Protein p53; Proteins; Regulation; Research; Risk; Role; Sampling; Serine; Signal Pathway; Signal Transduction; Signal Transduction Pathway; Specimen; Stem cells; Stress; TP53 gene; Testing; Transgenic Mice; Translations; Viral; Virus Diseases; Withdrawal; Work; base; cell growth; chemotherapy; cytokine; deprivation; eIF-2 Kinase; gene discovery; high risk; insight; leukemia; mouse model; mutant; novel; novel therapeutics; progenitor; public health relevance; response; small molecule; tumorigenesis

DESCRIPTION (provided by applicant): Interleukin 3 (IL-3) is a multi-potential hematopoietic growth factor that activates cellular signaling pathways to mediate growth and differentiation of bone marrow progenitor cells. Removal of IL-3 from factor- dependent cells induces cell cycle arrest and apoptosis. Our laboratory has discovered that cellular stresses, including IL-3 withdrawal from factor-dependent hematopoietic cells, inflammatory cytokines, chemotherapy treatment or viral infection promotes activation of the double-stranded RNA dependent protein kinase, PKR, by its cellular activator protein, RAX. Once activated, PKR inhibits protein synthesis by phosphorylating the alpha subunit of eIF2 to initiate apoptosis. Significantly, our findings reveal that RAX phosphorylation on serine 18 is required for PKR activation, translation inhibition and apoptosis initiated by diverse cellular stresses. In addition, we identified a novel RAX-dependent mechanism for regulation of tumor suppressor p53 transcriptional activity during cellular stress and discovered that reduced levels of endogenous RAX or forced expression of a dominant negative RAX mutant will promote aberrant cell growth. Significantly, our preliminary data indicate that PKR is differentially localized and activated in IPSS high risk compared to low risk MDS patient samples or normal hematopoietic progenitors. Taken together these findings suggest that the RAX- PKR stress signaling pathway may be critical for maintaining the correct composition of bone marrow cells and for initiating the response to infection from foreign agents. We hypothesize that PKR is activated by RAX during stress to promote both eIF2 phosphorylation and p53 activation that leads to translation inhibition, changes in gene expression, cell cycle arrest and apoptosis. Thus, inhibition or activation of PKR activity may be therapeutically useful in certain conditions. Specifically, PKR activators may be useful for promoting apoptosis of leukemic cells that have reduced levels of PKR, while small molecules that can inhibit PKR activity may be useful for treating hematopoietic disorders that display increased levels of activated PKR associated with aberrant apoptosis of bone marrow progenitor cells, such as in Fanconi anemia (FA) and Myelodysplastic Syndrome (MDS) that progress to AML. To test these hypotheses we will 1) determine the mechanism(s) by which RAX-PKR signal transduction regulates the cell cycle and initiates apoptosis following cellular stress in hematopoietic cells and 2) determine the significance of PKR in MDS progression to acute leukemia. For these studies we will use molecular biochemical and cell culture approaches and make use of the NUP98- HOXD13 transgenic mouse model that faithfully mimics MDS and progression to AML observed in humans. By achieving these specific aims we will gain new and important insights that fill knowledge gaps about how cellular stress activates the RAX-PKR signaling axis to regulate cell growth and apoptosis. A novel therapeutic strategy for treating patients with MDS and AML is expected to be the outcome. PUBLIC HEALTH RELEVANCE: The results generated by this research will increase our understanding of both the molecular mechanism(s) by which PKR promotes the cellular stress response and PKR's role in MDS evolution to AML. Results will point the way to and provide a basis for discovering novel anti-leukemia therapies.

描述（由申请人提供）：白细胞介素3（IL-3）是一种多潜能造血生长因子，可激活细胞信号传导途径以介导骨髓祖细胞的生长和分化。从因子依赖性细胞中去除IL-3诱导细胞周期停滞和凋亡。我们的实验室已经发现，细胞应激，包括IL-3从因子依赖性造血细胞中撤出、炎性细胞因子、化疗治疗或病毒感染，通过其细胞活化蛋白RAX促进双链RNA依赖性蛋白激酶PKR的活化。一旦被激活，PKR通过磷酸化eIF 2的α亚基以启动凋亡来抑制蛋白质合成。值得注意的是，我们的研究结果表明，丝氨酸18上的RAX磷酸化是PKR激活，翻译抑制和细胞凋亡所需的各种细胞应激。此外，我们确定了一种新的RAX依赖性机制，在细胞应激过程中调节肿瘤抑制因子p53的转录活性，并发现内源性RAX水平降低或显性负性RAX突变体的强制表达将促进异常细胞生长。值得注意的是，我们的初步数据表明，PKR是差异定位和激活IPSS高风险相比，低风险MDS患者样本或正常造血祖细胞。总之，这些发现表明RAX-PKR应激信号传导途径可能对于维持骨髓细胞的正确组成和启动对外来因子感染的应答至关重要。我们假设PKR在应激过程中被RAX激活，以促进eIF 2磷酸化和p53激活，从而导致翻译抑制、基因表达变化、细胞周期阻滞和凋亡。因此，PKR活性的抑制或激活在某些病症中可能是治疗上有用的。具体地，PKR激活剂可用于促进具有降低的PKR水平的白血病细胞的凋亡，而可抑制PKR活性的小分子可用于治疗表现出与骨髓祖细胞的异常凋亡相关的激活的PKR水平增加的造血疾病，例如发展为AML的范可尼贫血（FA）和骨髓增生异常综合征（MDS）。为了检验这些假设，我们将1）确定RAX-PKR信号转导调节细胞周期和在造血细胞中细胞应激后启动细胞凋亡的机制，和2）确定PKR在MDS进展为急性白血病中的意义。对于这些研究，我们将使用分子生物化学和细胞培养方法，并利用NUP 98-HOXD 13转基因小鼠模型，该模型忠实地模拟了在人类中观察到的MDS和AML进展。通过实现这些具体目标，我们将获得新的和重要的见解，填补有关细胞应激如何激活RAX-PKR信号传导轴以调节细胞生长和凋亡的知识空白。一种新的治疗策略，治疗MDS和AML患者的预期结果。公共卫生关系：这项研究的结果将增加我们对PKR促进细胞应激反应的分子机制和PKR在MDS向AML演变中的作用的理解。结果将为发现新的抗白血病疗法指明方向并提供依据。