In vivo modification of major histocompatibility complex class II DRA promoter occupancy mediated by the AIR-1 trans-activator.
RJ 2.2.5 is a human B cell mutant derived from the Burkitt lymphoma Raji cell which is defective in the AIR-1 locus function. This locus encodes a transcriptional trans-activator required for the constitutive expression of major histocompatibility complex (MHC) class II genes. Here we show, by in vivo DNase I footprinting, that the AIR-1 locus defect correlates with changes in the DRA promoter occupancy. Interestingly, reexpression of human MHC class II genes in RJ 2.2.5 x mouse spleen cell hybrids is associated with partial reversion of DRA promoter occupancy to the Raji pattern. DRA promoter occupancy in other class II-negative B cell lines, derived from patients with bare lymphocyte syndrome, is drastically different from the one observed in RJ 2.2.5 and Raji cells. Moreover, the use of the DNase I as an in vivo footprinting agent reveals that the patients' cell lines do not display a completely "bare promoter" as previously reported using dimethyl sulfate as the footprinting agent. Thus, the use of DNase I allowed us, for the first time, to correlate the AIR-1 locus defect with class II promoter occupancy alterations and distinguish these alterations from the ones observed in phenotypically similar but genetically distinct MHC class II-negative cells.
Rigaud
G
G
Institute of Immunology and Infectious Diseases, School of Medicine, University of Verona, Italy.
Paiola
F
F
Accolla
R S
RS
D001483:Base Sequence; D001665:Binding Sites; D017931:DNA Primers / Q000737:chemistry; D004268:DNA-Binding Proteins / Q000378:metabolism*; D003850:Deoxyribonuclease I / Q000378:metabolism; D006684:HLA-DR Antigens / Q000235:genetics*; D059805:HLA-DR alpha-Chains; D006801:Humans; D066298:In Vitro Techniques; D008214:Lymphocytes / Q000378:metabolism*; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D013463:Sulfuric Acid Esters / Q000737:chemistry; D015534:Trans-Activators / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D017931:DNA Primers; D004268:DNA-Binding Proteins; D006684:HLA-DR Antigens; D059805:HLA-DR alpha-Chains; D013463:Sulfuric Acid Esters; D015534:Trans-Activators; D003850:Deoxyribonuclease I; C007482:dimethyl sulfate