The Role of Zinc Finger Genes in Leukemogenesis

描述（由申请人提供）：锌指基因在白血病发生中的作用：拟议研究的目的是通过鉴定Ikaros蛋白的生物学功能来了解正常造血和造血细胞的恶性转化。Ikaros是正常造血和T细胞分化所必需的，并作为肿瘤抑制因子。Ikaros基因交替剪接产生多个锌指蛋白，参与基因调控和染色质重塑。我们的初步数据表明：1)在细胞周期的G1和S期，Ikaros在多个进化上保守的位点被CK2和其他激酶磷酸化；2)Ikaros特定氨基酸的磷酸化调节其dna结合能力和亚细胞定位。3) Ikaros在体内与Bcl-xL基因启动子结合，Ikaros活性失调与Bcl-xL基因表达上调有关。具体目标1：确定位点特异性ck2介导的磷酸化如何通过控制其作为分化和细胞周期进程调节剂的功能来促进Ikaros的肿瘤抑制活性。我们假设Ikaros的磷酸化会干扰其在转录调控和染色质重塑中的功能，并影响细胞增殖。我们将明确CK2靶位点Ikaros磷酸化在Ikaros靶基因染色质重塑中的作用。Ikaros的磷酸化在体内调节T细胞分化和T细胞增殖中的作用将被研究。靶向破坏Ikaros的小鼠的小鼠干细胞将被含有野生型Ikaros或拟磷Ikaros突变体的逆转录病毒载体感染，并移植到亚致死辐照的Ikaros敲除小鼠中。拟磷Ikaros突变体恢复正常T细胞分化的能力将与野生型Ikaros进行比较。具体目的#2：剖析CK2- Ikaros介导的Bcl-xL表达调控机制及其对Ikaros肿瘤抑制功能的贡献。先前的研究表明Ikaros可以抑制Bcl-xL基因的表达。我们假设Ikaros对Bcl-xL转录的负调控有助于Ikaros作为肿瘤抑制因子的功能。我们将研究CK2激酶介导的Ikaros磷酸化在Bcl-xL转录调控中的作用。我们将以靶向破坏Ikaros的小鼠为模型，在体内测试Bcl-xL的组成表达是否干扰Ikaros的抑瘤作用。这些研究将首次提供控制Ikaros肿瘤抑制功能的信号转导途径的详细功能分析。我们的研究将为控制造血细胞增殖的机制提供新的和重要的信息，并将为白血病的病理生理和治疗提供见解。公共卫生相关性：这项拟议研究的目的是了解正常血液形成和免疫系统发育的机制，以及导致白血病发展的变化。该项目的目标是确定Ikaros基因的功能。ikros的正常功能对正常的血液形成和免疫系统的发育至关重要。Ikaros基因活性的改变与儿童急性淋巴细胞白血病（ALL）、婴儿白血病和青少年慢性淋巴细胞白血病的发展有关。因此，我们的结果将有助于深入了解正常血液形成和白血病发展的机制。进一步阐明恶性转化过程的机制将有助于为白血病/淋巴瘤患者提供新的和更有效的治疗选择。

DESCRIPTION (provided by applicant): The Role of Zinc Finger Genes in Leukemogenesis: The objective of the proposed study is to understand normal hematopoiesis and the malignant transformation of hematopoietic cells by identifying the biological functions of Ikaros proteins. Ikaros is essential for normal hematopoiesis and T cell differentiation and acts as a tumor suppressor. The Ikaros gene is alternately spliced to generate multiple zinc finger proteins involved in gene regulation and chromatin remodeling. Our preliminary data show that: 1) Ikaros is phosphorylated at multiple evolutionarily conserved sites by CK2 and other kinases during G1 and S phase of the cell cycle 2) Phosphorylation of Ikaros at specific amino acids regulates its DNA-binding ability and subcellular localization. 3) Ikaros binds to the Bcl-xL gene promoter in vivo and disregulation of Ikaros activity is associated with upregulation of Bcl-xL gene expression. The specific aims of our proposals are: Specific Aim #1: Determine how site-specific CK2-mediated phosphorylation contributes to Ikaros' tumor suppressor activity by controlling its function as a regulator of differentiation and cell cycle progression. We hypothesize that phosphorylation of Ikaros interferes with its function in transcriptional regulation and chromatin remodeling and influences cellular proliferation. We will define the role of Ikaros phosphorylation at CK2 target sites in the chromatin remodeling of Ikaros target genes. The role of Ikaros' phosphorylation in regulating T cell differentiation and T cell proliferation will be studied in vivo. Murine stem cells from mice with targeted disruption of Ikaros will be infected with retroviral vectors containing wild type Ikaros or phosphomimetic Ikaros mutants and transplanted into sublethally irradiated Ikaros knockout mice. The ability of phosphomimetic Ikaros mutants to restore normal T cell differentiation will be compared to that of wild type Ikaros. Specific aim #2: Dissect the mechanism of CK2- Ikaros-mediated regulation of Bcl-xL expression and its contribution to Ikaros function as a tumor suppressor. Previous studies suggest that Ikaros represses Bcl-xL gene expression. We hypothesize that the negative regulation of Bcl-xL transcription by Ikaros contributes to Ikaros' function as a tumor suppressor. The role of CK2 kinase-mediated phosphorylation of Ikaros in regulation of Bcl-xL transcription will be studied. We will test whether constitutive expression of Bcl-xL interferes with Ikaros' tumor suppression in vivo using mice with targeted disruption of Ikaros as a model. These studies will provide the first detailed functional analysis of the signal transduction pathways that control the tumor suppressor function of Ikaros. Our research will provide new and important information on the mechanisms controlling the proliferation of hematopoietic cells and will yield insights into the pathophysiology and treatment of leukemia. PUBLIC HEALTH RELEVANCE: The objective of the proposed study is to understand the mechanism of normal blood formation and immune system development, and changes that lead to development of leukemia. The goal of the proposed project is to identify the function of Ikaros gene. Normal function of Ikaros is essential for normal blood forming and immune system development. Altered activity of the Ikaros gene is associated with the development of childhood acute lymphoblastic leukemia (ALL), infant leukemia and juvenile chronic lymphocytic leukemia. Thus, our results will help to gain insights into the mechanism of normal blood forming and development of leukemia. Further elucidation of the mechanism of the malignant transformation process will aid in providing novel and more effective treatment options for patients with leukemia/lymphoma.