Two distinct nuclear factors bind the conserved regulatory sequences of a rabbit major histocompatibility complex class II gene.
The constitutive coexpression of the major histocompatibility complex (MHC) class II genes in B lymphocytes requires positive, trans-acting transcriptional factors. The need for these trans-acting factors has been suggested by the reversion of the MHC class II-negative phenotype of rare B-lymphocyte mutants through somatic cell fusion with B cells or T-cell lines. The mechanism by which the trans-acting factors exert their effect on gene transcription is unknown. The possibility that two highly conserved DNA sequences, located 90 to 100 base pairs (bp) (the A sequence) and 60 to 70 bp (the B sequence) upstream of the transcription start site of the class II genes, are recognized by the trans-acting factors was investigated in this study. By using the gel electrophoresis retardation assay, a minimum of two proteins which specifically bound the conserved A or B sequence of a rabbit DP beta gene were identified in murine nuclear extracts of a B-lymphoma cell line, A20-2J. Fractionation of nuclear extract through a heparin-agarose column allowed the identification of one protein, designated NF-MHCIIB, which bound an oligonucleotide containing the B sequence and protected the entire B sequence in the DNase I protection analysis. Another protein, designated NF-MHCIIA, which bound an oligonucleotide containing the A sequence and partially protected the 3' half of this sequence, was also identified. NF-MHCIIB did not protect a CCAAT sequence located 17 bp downstream of the B sequence. The possible relationship between these DNA-binding factors and the trans-acting factors identified in the cell fusion experiments is discussed.
D000818:Animals; D001402:B-Lymphocytes / Q000378:metabolism; D001483:Base Sequence; D004247:DNA / Q000378:metabolism*; D005802:Genes, MHC Class II; D006367:HeLa Cells / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D011817:Rabbits; D013601:T-Lymphocytes / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*
D014157:Transcription Factors; D004247:DNA