CHIMERIC MRNA TEMPLATES DRIVE AIDS B CELL LYMPHOMAS

这一提议将检验一个与以下机制有关的独特假设 在B细胞恶性肿瘤中观察到的染色体易位 尤其与艾滋病淋巴瘤有关。我们建议嵌合体 免疫球蛋白(Ig)：癌基因信使核糖核酸在细胞间转录形成 初级RNA，诱导免疫球蛋白和免疫球蛋白之间的“不合理的同型转换” 癌基因位点，导致一种特殊形式的Ig：C-myc染色体 易位。我们将直接测试我们假设的中心原则-- 这种嵌合的mRNA(例如，IgH：C-myc)驱动了IgH癌基因的易位。目标 #1测试嵌合基因是否强制表达“桥接模板(S)” 在体外，会导致IgH-myc易位。小鼠B细胞系(I-29)， 已知在内毒素或转化生长因子-β刺激下转换为IgA，将被感染 (使用腺病毒载体)和适当的IgA：C-myc结构(S) 表达IgA：C-myc嵌合mRNA(Ia：C-myc外显子2和/或c- MycExon1：CA1)。细胞将被刺激进行IGH交换 预测这将导致IGA：C-myc的出现 重新安排。原代小鼠和人类B细胞也将感染 相应的a：c-myc或u：c-myc结构。单元格 也被刺激进行Ig同型转换和出现 IgH：评估C-myc易位。在这两种方法中， 将对易位进行测量，并确定其确切性质。目标2 检测Ig：C-myc嵌合mRNA是否“桥接” 模板(S)：体外结果免疫球蛋白：c-myc易位与肿瘤 发展。BALB/c AN小鼠将用同基因B细胞重建 携带：c myc构造，可导致：c-myc在 目标1.这种细胞的转移被预测会导致更快和更快的 浆细胞瘤的频繁发展。SCID小鼠将被重组 人类扁桃体B细胞携带u：c myc结构，驱动u：c myc 重排(目标1)。这种细胞的转移被预测为导致 人u：c-myc淋巴瘤在SCID小鼠体内的发生。AIM#3测试 嵌合的IgH：c-myc的mRNA，当作为转基因表达在 体内，驱动IgH：c-myc IgH：c myc mRNA的，当作为转基因表达在 体内，推动IgH：C-myc在B细胞中的易位并导致B细胞增强 细胞肿瘤的发生。编码一个或一对iga：c myc的构建体 嵌合mRNA将作为转基因导入Balb/c小鼠或将 作为转基因基因被引入ES细胞，将用于 重组RAG2基因缺陷小鼠。免疫球蛋白：C-myc的研究进展 易位和淋巴瘤将在这两个模型中进行评估， 免疫球蛋白A：C-myc对早期和多发淋巴瘤的预测 易位。

This proposal will test a unique hypothesis pertaining to the mechanism of chromosomal translocations observed in B cell malignancies that are particularly relevant in AIDS lymphomas. We propose that chimeric immunoglobulin (Ig): oncogene mRNA's formed by transplicing between the primary RNAs, induce "illegitimate isotype switching" between the Ig and oncogene loci, resulting in a specific form of Ig:c-myc chromosomal translocation. We will directly test the central tenet of our hypothesis-- that chimeric mRNA (e.g. IgH:c-myc) drive IgH-oncogene translocations. Aim #1 tests whether forced expression of chimeric mRNA "bridging template(s)" in vitro results in IgH-myc translocations. A murine B cell line (I-29), known to switch to IgA under LPS or TGF-beta stimulation, will be infected (using adenovirus vectors) with the appropriate Iga:c-myc construct(s) that express Iga:c-myc chimeric mRNA (Ia:c-myc Exon2 and/or c- mycExon1:Ca1). Cells will be stimulated to undergo IgH switching with the prediction that this will result in the appearance of Iga:c-myc rearrangements. Primary mouse and human B cells will also be infected with the appropriate a:c-myc or u:c-myc constructs respectively. Cells will also be stimulated to undergo Ig isotype switching and the appearance of IgH:c-myc translocations assessed. In both approaches, the number of translocations will be measured and their exact nature determined. AIM #2 tests whether introduction of Ig:c-myc chimeric mRNA "bridging template(s)" in vitro results in vivo IgH:c- myc translocations and tumor development. Balb/c AN mice will be reconstituted with syngeneic B cells carrying a:c myc constructs shown to result in a:c-myc rearrangement in Aim 1. Transfer of such cells is predicted to result in the more rapid and frequent development of plasmacytomas. SCID mice will be reconstituted with human tonsil B cells carrying u: c myc constructs that drive u: c myc rearrangement (Aim 1). Transfer of such cell is predicted to result in the development of u:c myc human lymphomas in the SCID mice. Aim #3 tests whether the chimeric IgH:c myc mRNA's, when expressed as transgenes in vivo, drive IgH:c-myc IgH:c myc mRNA's, when expressed as transgenes in vivo, drive IgH:c-myc translocations in B cells and result in enhanced B cell tumor development. Constructs encoding one or the pair of Iga:c myc chimeric mRNA's will be introduced as a transgene into Balb/c mice or will be introduced as a transgene into ES cells that will be use to reconstitute RAG 2 deficient mice. The development of Iga:c-myc translocations and lymphomas will be assessed in both models with the prediction of earlier and more frequent lymphomas being Iga:c-myc translocations.