A V(D)J RECOMBINASE-INDUCIBLE B-CELL LINE - ROLE OF TRANSCRIPTIONAL ENHANCER ELEMENTS IN DIRECTING V(D)J RECOMBINATION

Rapid analysis of mechanisms that regulate V(D)J recombination has bee n hampered by the lack of appropriate cell systems that reproduce aspe cts of normal prelymphocyte physiology in which the recombinase is act ivated, accessible antigen receptor loci are rearranged, and rearrange ment status is fixed by termination of recombinase expression. To gene rate such a system, we introduced heat shock-inducible V(D)J recombina tion-activating genes (RAG) 1 and 2 into a recombinationally inert B-c ell line. Heat shock treatment of these cells rapidly induced high lev els of RAG transcripts and RAG proteins that were accompanied by a par allel induction of V(D)J recombinase activity, strongly suggesting tha t RAG proteins have a primary role in V(D)J recombination. Within hour s after induction, these cells began to rearrange chromosomally integr ated V(D)J recombination substrates but only if the substrates contain ed an active transcriptional enhancer; substrates lacking an enhancer were not efficiently rearranged. Activities necessary to target integr ated substrates for rearrangement were provided by two separate lympho id-specific transcriptional enhancers, as well as an active nonlymphoi d enhancer, unequivocally demonstrating that such elements enhance bot h transcription and V(D)J recombinational accessibility.