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A critical role for C/EBPβ binding to the AABS promoter response element in the human iNOS gene
Author(s) -
Guo Zhong,
Shao Lifang,
Feng Xuesheng,
Reid Kaye,
Marderstein Eric,
Nakao Atsunori,
Geller David A.
Publication year - 2003
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fj.02-1172fje
Subject(s) - microbiology and biotechnology , promoter , response element , transcription factor , biology , ccaat enhancer binding proteins , gene , activator (genetics) , liver cell , binding site , hepatic stellate cell , gene expression , chemistry , dna binding protein , biochemistry , medicine , endocrinology
The human iNOS (hiNOS) gene is expressed in a tissue‐specific manner, but the molecular basis for this regulation has not been elucidated. Here, we show that liver cell‐specific hiNOS gene activation involves protein‐DNA binding to an A‐activator binding site (AABS) located at ‐192 nucleotides in the hiNOS promoter region. Mutation of this site in the −7.2 kb hiNOS promoter construct inhibited basal hiNOS promoter activity in primary rat hepatocytes (77%), and two human liver cell lines, AKN‐1 (63%) and HepG2 (60%), but had no significant effect on basal hiNOS activity in three non‐hepatic human cell types. Interestingly, mutation of AABS significantly abrogated cytokine‐induced promoter activity in all cell types. C/EBPβ transcription factor bound to AABS by gel shift assay. Overexpression of C/EBPβ active form (LAP) increased hiNOS basal promoter activity ∼sixfold in liver cells, but had minimal effect in non‐hepatic cells. In contrast, overexpression of the transcriptional inhibitor (LIP) strongly suppressed both basal and cytokine‐inducible promoter activity. These data show that the cis‐acting AABS DNA element mediates liver‐specific basal hiNOS promoter activity through binding of the trans‐acting C/EBPβ factor. Further, C/EBPβ binding to AABS functions as a “switchpoint” that is necessary for cytokine‐inducible hiNOS gene expression in all cell types examined.