NF-HB (BSAP) is a repressor of the murine immunoglobulin heavy-chain 3' alpha 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' alpha enhancer (3' alpha E). These sites were defined by oligonucleotide competition in an electrophoretic mobility shift assay (EMSA) and methylation interference footprinting. The 3' alpha E-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' alpha E-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' alpha E is one of the mammalian targets for NF-HB (BSAP). Transient-transfection assays with chloramphenicol acetyltransferase gene constructs containing 3' alpha E and mutant 3' alpha E, 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' alpha E over parental 3' alpha E 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 repressor of the mouse IgH 3' alpha E.
Department of Cell Biology, Albert Einstein College of Medicine, Bronx, New York 10461.
M
M
Singh
B K
BK
Birshtein
D000818:Animals; D001402:B-Lymphocytes / Q000166:cytology / Q000276:immunology*; D001483:Base Sequence; D001665:Binding Sites; D002454:Cell Differentiation; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004742:Enhancer Elements, Genetic; D005803:Genes, Immunoglobulin; D007143:Immunoglobulin Heavy Chains / Q000235:genetics*; D016393:Lymphoma, B-Cell; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009687:Nuclear Proteins / Q000378:metabolism*; D009838:Oligodeoxyribonucleotides / Q000378:metabolism; D051757:PAX5 Transcription Factor; D010954:Plasmacytoma; D011994:Recombinant Proteins / Q000378:metabolism; D012097:Repressor Proteins / Q000378:metabolism*; D015183:Restriction Mapping; D014157:Transcription Factors / Q000378:metabolism; D014162:Transfection; D014407:Tumor Cells, Cultured; D014466:Ultraviolet Rays
D004268:DNA-Binding Proteins; D007143:Immunoglobulin Heavy Chains; D009687:Nuclear Proteins; D009838:Oligodeoxyribonucleotides; D051757:PAX5 Transcription Factor; C497282:Pax5 protein, mouse; D011994:Recombinant Proteins; D012097:Repressor Proteins; D014157:Transcription Factors; D015500:Chloramphenicol O-Acetyltransferase