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.
