INVOLVEMENT OF THE EVI-2 LOCUS IN MYELOID LEUKEMIA

亲水性病毒整合位点-2(Evi-2)是一种常见的 BXH-2小鼠髓系肿瘤中病毒整合的部位。 对EVI-2基因座的鉴定表明，病毒整合 发生在2个编码跨膜蛋白的新基因(Evi-2a和Evi-2a EVI-2b)。EVI-2定位于小鼠11号染色体和人类 染色体17q11.2。通过使用EVI-2，NF1的基因被 克隆了EVI-2a、EVI-2b和第三个基因(Omgp)。 定位于内含子内。有趣的是，人们观察到 NF1患者有更高的青少年发育率 髓系白血病，表明无论是什么基因参与了 小鼠髓系白血病，可能与人类髓系白血病基因相同 白血病。因此，通过使用共同的病毒整合部位 发现了一个重要的基因组区域，该区域与小鼠和 人类疾病。 我们的长期目标是描述在人类基因组中发现的 EVI-2基因座并确定其在正常发育和 白血病。初步分析表明，不是这一变化。 NF1基因的表达，但基因(或多个基因)的改变 在导致髓系白血病的NF1内含子中。BXH-2小鼠 将老化以隔离更多的髓系肿瘤和内部整合 将确定EVI-2基因座。这些肿瘤将在 RNA和蛋白质水平来检测表达的变化 EVI-2基因座内的基因。此外，我们建议 鉴定EVI-2a和EVI-2b基因及其产物以确定 它们在正常发育和髓系白血病发生中的作用。完整的 这两个基因的长度mrna将被测序，其特征是 外显子的拼接模式和描绘。这些蛋白质将是 在表达载体中产生并用于产生多克隆抗血清 研究蛋白质在细胞内的定位。对这一事件的调查 蛋白质的功能将在鉴定的同时启动 相关联的蛋白质。此外，一个发展的概况， 用原位RNA杂交法同时表达mRNA和蛋白质 并进行免疫组织化学染色，以获得 洞察这些基因在小鼠正常发育中所起的作用。这个 这些基因产物的过度表达和缺失的影响 将在体外进行评估，以及过度表达对 转基因动物。这些研究将阐明细胞的正常功能。 EVI-2基因座及其在髓系肿瘤中的作用

Ecotropic viral integration site-2 (Evi-2) was identified as a common site of viral integration in myeloid tumors derived from BXH-2 mice. Characterization of the Evi-2 locus revealed that the viral integrations occurred within 2 novel genes encoding transmembrane proteins (Evi-2a and Evi-2b). Evi-2 was localized to mouse chromosome 11 and to human chromosome 17q11.2. Through the use of Evi-2, the gene for NF1 was cloned and it was revealed that Evi-2a, Evi-2b and a third gene (Omgp) were localized within an intron. Interestingly, it has been observed that patients with NF1 have a higher frequency of developing juvenile myelogenous leukemia, suggesting that whatever gene is involved in the murine myeloid leukemia, could be the same gene involved in human myeloid leukemia. Thus, through the use of a common site of viral integration an important genomic region was identified that is involved in murine and human disease. Our long term goal is to characterize the novel genes found within the Evi-2 locus and to determine their role in normal development and leukemia. Preliminary analysis indicates that it is not the alteration of expression of the NF1 gene, but the alteration of the gene (or genes) in the NF1 intron that contributes to the myeloid leukemia. BXH-2 mice will be aged to isolate additional myeloid tumors and integrations within the Evi-2 locus will be identified. These tumors will be analyzed at both the RNA and protein level to detect alterations in the expression of the genes within the Evi-2 locus. Additionally, we propose to characterize the Evi-2a and Evi-2b genes and their products to determine their role in normal development and myeloid leukemogenesis. The full length mRNA of both genes will be sequenced with the characterization of splicing patterns and delineation of exons. The proteins will be produced in expression vectors and used to generate polyclonal antisera to study the cellular localization of the proteins. Investigation of the function of the proteins will be initiated along with the identification of associated proteins. Additionally, a developmental profile of the expression of both mRNA and protein, through in situ RNA hybridization and immunohistochemical staining, will be determined in order to gain insights into the role these genes play in normal mouse development. The effect of overexpression and loss of expression of these gene products in vitro will be assessed as well as the effect of overexpression in transgenic animals. These studies will elucidate the normal function of the Evi-2 locus and its influence in myeloid neoplasia.