RETINOIC ACID RECEPTOR A IN GRANULOPOIESIS AND LEUKEMIA

全反式维甲酸促进几种中性粒细胞分化 并诱导急性髓系白血病患者完全缓解 早幼粒白血病(APL)。在APL中，维甲酸受体基因 (RAR-α)被转移到新的位点PML，导致 融合基因PML-RAR-α的构建及表达。然而， RAR-α和PML-RAR-α在骨肉瘤发生发展中的确切作用 中性粒细胞和白血病的关系尚不清楚。这项建议的具体目的 主要有：(1)测定维甲酸(RA)和维甲酸 受体-α(RAR-α)在分化过程中起重要作用 中性粒细胞；(2)调查 融合基因PML-RAR-α在髓系细胞中的表达 将使用三种方法来确定RA和RAR-α是否在 在中性粒细胞分化中起着至关重要的作用。首先，我们将 利用逆转录病毒载体表达“显性阴性”RAR-α 一种能够分化为中性粒细胞的造血细胞系 巨噬细胞。如果RAR-α对肿瘤的分化是必不可少的 中性粒细胞，显性负性突变体对其功能的抑制 应该会扰乱分化过程。第二，我们将使用相同的 转导和表达显性阴性的逆转录病毒载体 RAR-α在小鼠造血干细胞中的作用研究 体内的造血功能。第三，将使用无血清培养系统。 目的：确定中性粒细胞发育过程中RA的需要量。至 调查该基因表达的生物学后果 PML-RAR-α融合基因在髓系细胞中，我们将采用两种 战略。首先，携带融合基因的逆转录病毒载体将 用于感染具有中性粒细胞分化能力的髓系细胞 响应RA以确定该融合基因是否表现为显性 消极活动。第二，我们将感染小鼠骨髓干细胞 携带相同的逆转录病毒并将干细胞移植到接受者体内 研究融合基因对小鼠白血病的致病作用。 申请者的长期研究兴趣是分子控制 正常中性粒细胞和肿瘤中性粒细胞的分化。总而言之， 拟议的研究使我们对RA的角色有了更多的了解 和RAR-α在造血中的作用。

All-trans retinoic acid promotes neutrophilic differentiation of several myeloid cell lines and induces complete remission in patients with acute promyelocytic leukemia (APL). In APL, the gene of retinoic acid receptor (RAR-alpha) is translocated to a new locus PML, resulting in the generation and expression of a fusion gene PML-RAR-alpha. However, the exact roles of RAR-alpha and PML-RAR-alpha in the development of neutrophils and leukemia are unclear. The specific aims of this proposal are: (1) To determine If retinoic acid (RA) and retinoic acid receptor-alpha (RAR-alpha) play an essential role in the differentiation of neutrophils; (2) To investigate the biological consequences of the expression of the fusion gene PML-RAR-alpha in myeloid cells. Three approaches will be used to determine if RA and RAR-alpha play a crucial role in the differentiation of neutrophils. First, we will utilize a retroviral vector to express a "dominant negative" RAR-alpha in a hematopoietic cell line capable of differentiation into neutrophils and macrophages. If RAR-alpha is essential to the differentiation of neutrophils, suppression of its function by the dominant negative mutant should disrupt the differentiation process. Second, we will use the same retroviral vector to transduce and express the dominant negative RAR-alpha in mouse hematopoietic stem cells to study its effect on hematopoiesis in vivo. Third, a serum-free culture system will be used to determine the requirement of RA in the development of neutrophils. To investigate the biological consequences of the expression of PML-RAR-alpha fusion gene in myeloid cells, we will employ two strategies. First, a retroviral vector harboring the fusion gene will be used to infect myeloid cell lines capable of neutrophilic differentiation in response to RA to determine whether this fusion gene exhibits dominant negative activity. Second, we will infect murine bone marrow stem cells with the same retrovirus and transplant the stem cells into recipient mice to study the leukemogenic potential of the fusion gene. The applicant's long-term research interest is the molecular control of the differentiation of normal and neoplastic neutrophils. In summary, the proposed studies allow us to gain more insights into the roles of RA and RAR-alpha in hematopoiesis.