We investigated the cis-acting sequences that function in the B-cell-specific and interferon-gamma-inducible expression of the HLA-DR alpha gene, a human class II major histocompatibility complex gene. The effects of 5' deletions on the activity of the DR alpha promoter and the influence of upstream DR alpha promoter elements on the activity of the herpes simplex virus thymidine kinase promoter were examined by a transient transfection assay in human B-, T-, and fibroblast cell lines. We show that the DR alpha gene is regulated by positive and negative cis-acting sequences between positions -1300 and +31 from the site of initiation of transcription. We also demonstrate that the DR alpha promoter sequences from positions -116 to -92 and from -136 to -80 are the minimal sequences required for conferring B-cell specificity and interferon-gamma inducibility upon the Herpes simplex virus thymidine kinase promoter, respectively.