Transcriptional regulation of the p67phox gene: role of AP-1 in concert with myeloid-specific transcription factors.
We have investigated the myeloid-specific transcriptional regulation of p67(phox), an essential component of phagocyte respiratory burst NADPH oxidase. Analysis was carried out on the p67(phox) 5'-flanking region from -3669 to -4 (relative to ATG), including the first exon and intron and part of the second exon. The construct extending from -985 to -4 produced the highest luciferase activity in myeloid HL-60 cells but was not active in HeLa or Jurkat cells, indicating myeloid-specific expression. Four active elements were identified: Sp1/Sp3 at -694, PU.1 at -289, AP-1 at -210, and PU.1/HAF1 at -182, the latter three being in the first intron. These cis elements bound their cognate transacting factors both in vitro and in vivo. Mutation of the Sp1, PU.1, or PU.1/HAF1 site each decreased promoter activity by 35-50%. Mutations in all three sites reduced promoter activity by 90%. However, mutation of the AP-1 site alone nearly abolished promoter activity. The AP-1 site bound Jun and Fos proteins from HL-60 cell nuclear extract. Co-expression with Jun B in AP-1-deficient cells increased promoter activity by 3-fold. These data show that full p67(phox) promoter activity requires cooperation between myeloid-specific and nonmyeloid transcription factors, with AP-1 being the most critical for function.
Li
S L
SL
Department of Medicine, University of Texas Health Science Center and South Texas Veterans Health Care System, Audie L. Murphy Division, San Antonio, Texas 78229-3900, USA.
Valente
A J
AJ
Wang
L
L
Gamez
M J
MJ
Clark
R A
RA
D001483:Base Sequence; D001665:Binding Sites; D002467:Cell Nucleus / Q000378:metabolism; D003001:Cloning, Molecular; D004268:DNA-Binding Proteins / Q000378:metabolism; D003850:Deoxyribonuclease I / Q000378:metabolism; D015536:Down-Regulation; D005091:Exons; D015971:Gene Expression Regulation, Enzymologic; D018922:HL-60 Cells; D006367:HeLa Cells; D006801:Humans; D007438:Introns; D019169:Jurkat Cells; D008156:Luciferases / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010750:Phosphoproteins / Q000235:genetics* / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D016329:Sp1 Transcription Factor / Q000378:metabolism; D051705:Sp3 Transcription Factor; D018808:Transcription Factor AP-1 / Q000235:genetics* / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured
D004268:DNA-Binding Proteins; D010750:Phosphoproteins; C497121:SP3 protein, human; D016329:Sp1 Transcription Factor; D018808:Transcription Factor AP-1; D014157:Transcription Factors; C079789:neutrophil cytosol factor 67K; D051705:Sp3 Transcription Factor; D008156:Luciferases; D003850:Deoxyribonuclease I