Recognition and cleavage of cryptic recombination signal sequences identified from lymphoid malignancies (49.20)

Abstract

Abstract Immune repertoire diversity is largely achieved by V (D) J recombination, during which antigen receptor genes are assembled from arrays of component gene segments by site-specific DNA rearrangement. This process is initiated when the RAG proteins introduce DNA double-strand breaks (DSBs) at recombination signal sequences (RSSs) abutting antigen receptor coding segments. However, the RAG proteins occasionally mistarget DNA sequences outside of antigen receptor loci and promote chromosomal translocations and deletions, which are among the major events that initiate neoplasia in lymphoid organs. Previous studies have identified various RSS-like sequences involved in chromosomal translocations, including LMO2, TAL1, Ttg-1, and Hox11, and the interstitial deletion at 1p32 involving SIL/SCL. Here we have used electrophoretic mobility shift assays and in vitro cleavage assays to study RAG-mediated binding and cleavage to the oligonucleotide substrates containing these cryptic RSSs. We find that RAG proteins can cleave LMO2, TAL1, Ttg-1, and SIL in vitro, and introduce DSBs at these sites which can be detected by ligation-mediated PCR in cell culture. In contrast, Hox11 and SCL are nicked but fail to support DSB formation both in vitro and in cell culture. These data raise the possibility that Hox11 and SCL may be cleaved by an as-yet unidentified mechanism. This research is supported by American Cancer Society grant RSG-01-020-05-LIB to P.C.S.