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Identification of a classic cytokine‐inducible enhancer in the upstream human iNOS promoter
Author(s) -
Guo Zhong,
Shao Lifang,
Du Qiang,
Park Kyung Soo,
Geller David A
Publication year - 2006
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/fasebj.20.4.a511-b
Subject(s) - enhancer , promoter , microbiology and biotechnology , enhancer rnas , chromatin immunoprecipitation , biology , transcription factor , transcription (linguistics) , gene , gene expression , genetics , linguistics , philosophy
The human iNOS (hiNOS) promoter as far as 16 kb is one of the largest known promoters. Its transcriptional regulation is extremely complex involving the multiple pathways of cytokine signaling. We have previously reported that only hiNOS promoter beyond −3.8 kb showed any significant induction with cytokines mixture (CM) of TNFα, IL‐1β and IFN‐γ in human A549 and AKN 1cells. In this study, we further identified a classic enhancer located between −6 kb and −5 kb of hiNOS upstream promoter. By the analysis of serial constructs of hiNOS promoter, this important 1 kb promoter sequence functions as the cytokine‐inducible enhancer in an orientation‐ and position‐independent manner, and also conveys the same ability with the heterologous thymidine kinase (tk) promoter. Chromatin immunoprecipitation (ChIP) was applied to detect the in vivo protein/DNA interactions within this enhancer region. The functional bindings of NFκB and Stat‐1 at −5.8 kb human iNOS promoter site were significantly increased in A549 cells, while the binding of Stat‐1 not NFκB at −5.2 kb site was moderately increased. These results suggest that the −6 to −5 kb promoter region as a classic transcriptional enhancer for human iNOS gene can interact with specific NFκB and Stat‐1 nuclear proteins and enhance the transcription of hiNOS gene under cytokine stimulation.