: V(D)J recombination is normally associated with immunoglobulin and T cell receptor rearrangements. Aberrant genomic V(D)J-recombinase-mediated events are potential sources of oncogenic rearrangements. Many aspects of V(D)J recombination are shared by transposons, and V(D)J-mediated transposition has been demonstrated in vitro. Our hypothesis is that V(D)J-mediated transposition occurs in vivo and that genotoxic agents associated with tobacco smoke and dietary flavonoids lead to an increase in aberrant V(D)J recombination and transposition.
Specific Aim #1 To determine whether V(D)J-mediated transposition occurs in vivo. We will utilize a plasmid-based assay in an attempt to capture V(D)J-mediated transposition into the HPRTlocus. Secondly, clones from pediatric patients containing V(D)J-mediated deletion of HPRT exons 2 and 3 will be analyzed for exon 2 and 3 transposition.
Specific Aim #2 Using the plasmid-based assay above cells will be treated with genotoxic agents found in tobacco smoke and dietary flavonoids. This assay will allow us to assess the effects of genotoxic exposure on transpositional events as well as V(D)J-mediated aberrant recombination within the HPRT gene.
Specific Aim #3 Mechanistic studies will be initiated to examine the expression of known proteins associated with V(D)J recombination including Rag1, Rag2 and DNAPK. Altogether these experiments will help us correlate environmental exposure and the level of aberrant V(D)J recombination to the expression of known V(D)J components.
| Finette, Barry A (2006) Analysis of mutagenic V(D)J recombinase mediated mutations at the HPRT locus as an in vivo model for studying rearrangements with leukemogenic potential in children. DNA Repair (Amst) 5:1049-64 |
