Studies of expression of the DQ beta promoter and its 5' deletion derivatives in normal and mutant human B cell lines.
A series of 5' deletion test plasmids harboring promoter sequences of the HLA-DQ beta gene fused to the bacterial chloramphenicol acetyl transferase (CAT) gene were constructed. Transient CAT expression from these constructs in several types of cells was employed to examine the role of the promoter sequence in the regulation of DQ beta gene expression. The DQ beta constructs drove CAT expression in Raji cells (human Burkitt lymphoma cells) to at least 25-fold or 50-fold higher levels than in Hela cells (human cervical carcinoma cells) or Jurkat cells (human T-leukemia cells), respectively. A short promoter sequence of -160 bp containing the conserved X and Y sequences was sufficient for expression in Raji cells, and deletion to -106 bp which interrupted the X sequence abolished the expression. Sequences further upstream to -160 bp as far as -2500 bp which included an Ig-like octamer appeared to have no effect on expression in Raji cells. Thus, the promoter up to -160 bp has all of the sequences required for B cell specific expression of CAT in this assay. CAT expression from the 5' deletion constructs introduced into RJ2.2.5 and 6.1.6 cells (class II-negative mutant B cells lines) was also examined. None of the 5' deletion constructs, including that with -160 bp of promoter, showed CAT expression in these cells, suggesting that a transcriptional factor(s) required for the activity of the DQ beta promoter was missing in these cells. Moreover, the -160 to -66 bp sequence (including the X and Y elements), which functions as a B cell specific enhancer, was inactive in the mutant cells. Thus, the missing factor(s) are required for the enhancer function of this fragment.
Sakurai
M
M
Department of Biochemistry and Molecular Biology, Harvard University, Cambridge, MA.
Strominger
J L
JL
D001402:B-Lymphocytes / Q000276:immunology*; D001483:Base Sequence; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D005786:Gene Expression Regulation / Q000235:genetics; D005802:Genes, MHC Class II / Q000235:genetics; D006683:HLA-DQ Antigens / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D009154:Mutation; D010957:Plasmids; D011401:Promoter Regions, Genetic; D012045:Regulatory Sequences, Nucleic Acid; D013601:T-Lymphocytes / Q000276:immunology; D014157:Transcription Factors; D014162:Transfection
D006683:HLA-DQ Antigens; D014157:Transcription Factors; D015500:Chloramphenicol O-Acetyltransferase