PU.1 as an essential activator for the expression of gp91(phox) gene in human peripheral neutrophils, monocytes, and B lymphocytes.
We have reported a deficiency of a 91-kDa glycoprotein component of the phagocyte NADPH oxidase (gp91(phox)) in neutrophils, monocytes, and B lymphocytes of a patient with X chromosome-linked chronic granulomatous disease. Sequence analysis of his gp91(phox) gene revealed a single-base mutation (C --> T) at position -53. Electrophoresis mobility-shift assays showed that both PU.1 and hematopoietic-associated factor 1 (HAF-1) bound to the inverted PU.1 consensus sequence centered at position -53 of the gp91(phox) promoter, and the mutation at position -53 strongly inhibited the binding of both factors. It was also indicated that a mutation at position -50 strongly inhibited PU.1 binding but hardly inhibited HAF-1 binding, and a mutation at position -56 had an opposite binding specificity for these factors. In transient expression assay using HEL cells, which express PU.1 and HAF-1, the mutations at positions -53 and -50 significantly reduced the gp91(phox) promoter activity; however, the mutation at position -56 did not affect the promoter activity. In transient cotransfection study, PU.1 dramatically activated the gp91(phox) promoter in Jurkat T cells, which originally contained HAF-1 but not PU.1. In addition, the single-base mutation (C --> T) at position -52 that was identified in a patient with chronic granulomatous disease inhibited the binding of PU.1 to the promoter. We therefore conclude that PU.1 is an essential activator for the expression of gp91(phox) gene in human neutrophils, monocytes, and B lymphocytes.
Suzuki
S
S
Department of Biochemistry, Institute of Tropical Medicine, Nagasaki University, Nagasaki 852, Japan.
Kumatori
A
A
Haagen
I A
IA
Fujii
Y
Y
Sadat
M A
MA
Jun
H L
HL
Tsuji
Y
Y
Roos
D
D
Nakamura
M
M
D001402:B-Lymphocytes / Q000502:physiology*; D001665:Binding Sites / Q000235:genetics; D002675:Child, Preschool; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation; D006105:Granulomatous Disease, Chronic / Q000097:blood / Q000235:genetics*; D006367:HeLa Cells; D006609:High Mobility Group Proteins; D006801:Humans; D008297:Male; D008562:Membrane Glycoproteins / Q000096:biosynthesis / Q000235:genetics*; D008875:Middle Aged; D009000:Monocytes / Q000502:physiology*; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases / Q000235:genetics; D009504:Neutrophils / Q000502:physiology*; D011401:Promoter Regions, Genetic; D011518:Proto-Oncogene Proteins / Q000235:genetics* / Q000378:metabolism; D015534:Trans-Activators / Q000235:genetics* / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism; D000099232:Proto-Oncogene Protein Spi-1
D004268:DNA-Binding Proteins; D006609:High Mobility Group Proteins; D008562:Membrane Glycoproteins; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases; D011518:Proto-Oncogene Proteins; D015534:Trans-Activators; D014157:Transcription Factors; D000099232:Proto-Oncogene Protein Spi-1; C102078:HAF-1 protein, human; C056558:CYBB protein, human