CHIMERIC MRNA TEMPLATES DRIVE AIDS B CELL LYMPHOMAS

该提案将测试一个关于以下机制的独特假设： 在B细胞恶性肿瘤中观察到的染色体易位， 特别是在艾滋病淋巴瘤中。我们认为嵌合体 免疫球蛋白（IG）：癌基因mRNA的形成之间的转录 一级RNA诱导IG和 癌基因位点，导致特定形式的IG：c-myc染色体 易位我们将直接测试我们假设的核心原则 嵌合mRNA（例如IgH：c-myc）驱动IgH-癌基因易位。目的 #1测试嵌合mRNA "桥接模板"的强制表达是否 在体外导致IgH-myc易位。一种鼠B细胞系（I-29）， 已知在LPS或TGF-β刺激下转换为IgA，将被感染 （使用腺病毒载体）与适当的伊加：c-myc构建体 其表达伊加：c-myc嵌合mRNA（Ia：c-myc外显子2和/或c-myc外显子3）， mycExon1：Ca1）。 细胞将被刺激以经历IgH转换， 预测这将导致伊加：c-myc的出现 重新安排原代小鼠和人B细胞也将感染 分别是合适的a：c-myc或u：c-myc构建体。细胞将 也被刺激进行IG同种型转换， 评估IgH：c-myc易位。在这两种方法中， 将测量易位并确定其确切性质。目标#2 测试引入IG：c-myc嵌合mRNA "桥接"是否 模板"体外导致体内IgH：c-myc易位和肿瘤 发展BalB/c AN小鼠将用同基因B细胞重建 携带a：c-myc构建体的人显示导致a：c-myc重排， 目标1。预计这种细胞的转移将导致更快速和更有效的细胞转移。 浆细胞瘤的频繁发展。将重建SCID小鼠 人扁桃体B细胞携带驱动u：c myc的u：c myc结构， 重排（目的1）。这种细胞的转移预计会导致 在SCID小鼠中u：c myc人淋巴瘤的发展。目标#3测试 无论嵌合IgH：c myc mRNA在作为转基因表达时， 体内，驱动IgH：c-myc IgH：c myc mRNA，当作为转基因在 体内，驱动B细胞中的IgH：c-myc易位，并导致增强的B 细胞肿瘤的发展。编码一个或一对伊加：c myc的构建体 嵌合mRNA将作为转基因导入Balb/c小鼠或将 作为转基因导入ES细胞， 重组RAG 2缺陷型小鼠。伊加：c-myc基因的研究进展 易位和淋巴瘤将在两种模型中进行评估， 预测早期和更常见的淋巴瘤为伊加：c-myc 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.