MAPPING OF THE INTERACTION SITE OF THE DEFECTIVE TRANSCRIPTION FACTOR IN THE CLASS-II MAJOR HISTOCOMPATIBILITY COMPLEX MUTANT-CELL LINE CLONE-13 TO THE DIVERGENT X2-BOX

We have previously described a mutant B lymphoblastoid cell line, Clone-13, that expresses HLA-DQ in the absence of HLA-DR and -DP, Several criteria indicated that the defect in this cell line influences the activity of an isotype-specific transcription factor. Indeed, transient transfection of HLA-DRA and DQB reporter constructs indicated that the affected factor operates via cis-elements located between -141 base pairs and the transcription initiation site. A series of hybrid DRA/DQB reporter constructs was generated to further map the relevant cis-elements in this system, Insertion of oligonucleotides spanning the DQB X-box (but not the DQB-W region or the DQB Y-box) upstream of -141 in a DRA reporter plasmid rescued expression to nearly wild-type levels, Substitution promoters were then generated where the entire X-box, or only the X1- or X2-boxes of HLA-DRA were replaced with the analogous regions of HLA-DQB. The DQB X2-box was able to restore expression to the silent DRA reporter construct. Moreover, replacement of the DQB X2-box with the DRA X2-box markedly diminished the activity of the DQB promoter in the mutant cell, None of the hybrid reporter constructs were defective when transfected into the wild-type, HLA-DR/-DQ positive that the divergent X2-box of the class II major histocompatibility complex promoters plays an important role in influencing differential expression of the human class II isotypes.