NUCLEAR-PROTEIN BINDING TO OCTAMER MOTIFS IN THE IMMUNOGLOBULIN GAMMA-1 SWITCH REGION

Initiation of the immunoglobulin heavy chain switch DNA rearrangement event is thought to involve conversion of the target switch region DNA to an accessible state. Accessibility is likely to be mediated by the binding of regulatory proteins to sequences in or near switch regions. A DNase hypersensitivity assay was used to recognize possible regions of protein binding in the gamma-1 switch region of the B cell hybridoma 470.25. A strong DNase hypersensitive site was identified 5' of the tandemly repeated S-gamma-1 sequences. Data from other laboratories suggest that this hypersensitive site is associated with switch recombination to gamma-1. However, the 470.25 cell does not express the gamma-1 germline transcript. A second group of DNase hypersensitive sites within the repetitive portion of the gamma-1 switch region was also identified. A gel retardation assay for protein - DNA interaction revealed a sequence present in several copies in the gamma-1 switch region that specifically binds nuclear proteins. This binding sequence, SG1BS, contains the octanucleotide sequence ATGCAAAA, a 7/8 match to the transcriptional enhancer octamer motif found in immunoglobulin promoters and the heavy chain enhancer. Binding competition studies of SG1BS demonstrate that both the octamer and flanking sequences are critical for binding. By size- and tissue-distribution, the factors that bind SG1BS are not distinguishable from the previously identified octamer-binding factors OTF-1 and OTF-2. The ability of proteins to bind the S-gamma-1 octamer motif is increased 2.3-fold upon IL-4 induction of lipopolysaccharide-stimulated B cells.