Deletion of germline promoter PD beta 1 from the TCR beta locus causes hypermethylation that impairs D beta 1 recombination by multiple mechanisms

The role of the germline transcriptional promoter, PD beta1, in V(D)J recom bination at the T cell receptor beta locus was investigated. Deletion of PD beta1 caused reduced germline transcription and DNA hypermethylation in th e D beta1-J beta1 region and decreased D beta1 rearrangement Analyses of me thylation levels surrounding recombination signal sequences (RSS) before, d uring, and after recombination revealed that under physiological conditions cleavage of hypomethylated alleles was preferred over hypermethylated alle les. Methylation of a specific CpG site within the heptamer of the 3' D bet a1 RSS was incompatible with cleavage by the V(D)J recombinase. These findi ngs suggest that methylation can regulate V(D)J recombination both at a gen eral level by influencing regional chromatin accessibility and specifically by blocking RSS recognition or cleavage by the V(D)J recombinase.