The DNA-bending protein, HMG1, is required for correct cleavage of 23 bp recombination signal sequences by recombination activating gene proteins in vitro

DNA-bending proteins are known to facilitate the in vitro V(D)J joining of antigen receptor genes. Here we report that the high-mobility group protein , HMG1, is necessary for the correct nicking of the 23 bp recombination sig nal sequence (23-RSS) by the products of the recombination activating gene (RAG) proteins, RAG1 and RAGS. Without HMG1, the mouse J kappa 1 23-RSS was recognized as if it were the 12-RSS and nicked at a site 12 + 7 nucleotide s away from the 9mer signal, even though no 7mer-like sequence was evident at the cryptic nicking site. When increased amounts of HMG1 were added, the 23-RSS substrate was nicked correctly at a site 23 + 7 nucleotides from th e 9mer, and nicking at the cryptic site disappeared. Unlike the 23-RSS, the 12-RSS did not require HMG1 for correct nicking, although HMG1 was found t o increase the interaction between RSS and RAG proteins. Modification-inter ference assays demonstrated that HMG1 caused changes in the interaction bet ween the 23-RSS and RAG proteins specifically at the 7mer and the cryptic n icking site.