NF-HB (BSAP) is a repressor of the murine immunoglobulin heavy-chain 3′α enhancer at early stages of B-cell differentiation

We have identified a nuclear factor expressed in pro-B-, pre-B-, and B-cell lines that binds to two sites within the murine immunoglobulin heavy-chain (IgH) 3′α enhancer (3′αE). These sites were defined by oligonucleotide competition in an electrophoretic mobility shift assay (EMSA) and methylation interference footprinting. The 3′αE-binding factor is indistinguishable from NF-HB (B-lineage-specific nuclear factor that binds to the IgH gene) and the B-lineage-specific transcription factor BSAP by several criteria, including similar cell type distribution of binding activity, cross-competition of binding sites in EMSA, similar protein size as demonstrated by UV cross-linking, and sequence identity of one of the 3′αE-binding sites with a BSAP-binding site within the promoter of the sea urchin late histone gene H2A-2.2. These observations indicate that 3′αE is one of the mammalian targets for NF-HB (BSAP). Transient-transfection assays with chloramphenicol acetyltransferase gene constructs containing 3′αE and mutant 3′αE, in which one of the NF-HB binding sites was inactivated by site-specific mutagenesis, showed ca. five- to sixfold-enhanced activity of mutated 3′αE over parental 3′αE in B-cell lines (NF-HB⁺), while no significant difference was observed in plasmacytoma cells (NF-HB^-). We conclude from these observations that NF-HB (BSAP) acts as a represser of the mouse IgH 3′αE.