NPM-ALK AND ALK IN LYMPHOMA AND NORMAL DEVELOPMENT

描述：（改编自研究者摘要） 恶性肿瘤特异性染色体易位的表征 疾病导致了一些基因的鉴定， 在正常细胞发育或增殖中起关键作用， 当改变时产生异常生长。大约10%的非 霍奇金淋巴瘤具有t（2;5）（p23;q35）染色体重排。 在申请人实验室进行的定位克隆研究 揭示了t（2;5）产生了一个融合基因，该基因编码嵌合的 由核仁的氨基末端部分组成的蛋白质 磷蛋白核磷蛋白（NPM）连接到催化结构域的一个 胰岛素受体亚家族的新受体酪氨酸激酶， 间变性淋巴瘤激酶（ALK）。与ALK相互作用并 这种受体的正常功能是未知的。 因此 在NPM-ALK融合中，ALK激酶被组成性激活， 在淋巴样细胞中异位表达。此外，国家预防机制 嵌合体的残基与野生型NPM结合，靶向NPM的一部分。 融合蛋白到细胞核。目前尚不清楚， NPM-ALK的核定位是完整表达所必需的 它的生物活性。 本提案的长期目标是通过以下方式确定机制： NPM-ALK参与淋巴瘤发生，并确定其作用 ALK受体激酶在正常细胞生长和发育中的作用。的 实验计划是确定所需的结构基序， 激活ALK催化结构域和NPM-ALK核靶向， 以确定这些基序改变的生物学后果， 通过使用体外转化测定，以及基于 NPM-ALK嵌合体使造血细胞生长的能力 因子独立，以分析含有突变的构建体， 融合的NPM部分。嵌合蛋白的致瘤性 在生理条件下，将通过产生转基因 在启动子控制下表达NPM-ALK基因的小鼠 对淋巴细胞发育的不同阶段具有特异性。的作用 ALK在胚胎发育中的作用及其在特定组织中的正常功能 将通过三种互补的办法加以解决。一、格局 将在不同时间点的小鼠胚胎中测定ALK表达 发育阶段，并在选定的成人组织，通过原位 杂交和免疫细胞化学染色。第二，体内 将评估ALK受体失活的表型效应 通过有针对性地破坏小鼠的基因。三、表达克隆 策略将用于识别和表征同源配体 ALK的--决定ALK信号正常时的开关 发生。总之，这些研究应该提供一个机制， 关于t（2;5）激活ALK如何有助于 非霍奇金淋巴瘤的起源。

DESCRIPTION: (adapted from the investigator's abstract) The characterization of chromosomal translocations specific to malignant disease has led to the identification of a number of genes that play critical roles in normal cellular development or proliferation, and that produce abnormal growth when altered. Approximately 10% of all non- Hodgkin s lymphomas possess a t(2;5)(p23;q35) chromosomal rearrangement. Positional cloning studies performed in the applicants laboratory have revealed that the t(2;5) produces a fusion gene which encodes a chimeric protein consisting of the amino-terminal portion of the nucleolar phosphoprotein nucleophosmin (NPM) linked to the catalytic domain of a novel receptor tyrosine kinase of the insulin receptor subfamily, anaplastic lymphoma kinase (ALK). The ligand that interacts with ALK and the normal function(s) of this receptor are unknown. As a consequence of NPM-ALK fusion, the ALK kinase becomes constitutively activated and is expressed ectopically in lymphoid cells. In addition, the NPM residues of the chimera bind to wild-type NPM, targeting a portion of the fusion protein to the nucleus. It is presently unknown whether the nuclear localization of NPM-ALK is required for the complete expression of its biological activity. The long-range goals of this proposal are to determine the mechanisms by which NPM-ALK contributes to lymphomagenesis, and to determine the role of the ALK receptor kinase in normal cell growth and development. The experimental plan is to identify the structural motifs required for activation of the ALK catalytic domain and NPM-ALK nuclear targeting and to determine the biological consequences of alterations of these motifs by using in vitro transformation assays, as well as assays based on the ability of the NPM-ALK chimera to render hematopoietic cells growth factor-independent, to analyze constructs containing mutations of the NPM portion of the fusion. The oncogenicity of the chimeric protein under physiological conditions will be tested by generating transgenic mice that express the NPM-ALK gene under the control of promoters specific for different stages of lymphoid cell development. The role of ALK in embryonic development and its normal function in specific tissues will be addressed by three complementary approaches. First, the pattern of ALK expression will be determined in murine embryos at different developmental stages, and in selected adult tissues, by in situ hybridization and immunocytochemical staining. Second, the in vivo phenotypic effects of inactivation of the ALK receptor will be assessed by targeted disruption of the gene in mice. Third, expression cloning strategies will be used to identify and characterize the cognate ligand of ALK -- the on-off switch that determines when ALK signaling normally occurs. Taken together, these studies should provide a mechanistic framework for how activation of ALK by the t(2;5) contributes to the genesis of non-Hodgkin s lymphoma.