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Role of AP1 element in the activation of human eNOS promoter by lysophosphatidycholine
Author(s) -
Xing Feiyue,
Jiang Yong,
Liu Jing,
Zhao Kesen,
Mo Yongyan,
Qin Qinghe,
Wang Jingzhen,
Ouyang Jianming,
Zeng Yaoying
Publication year - 2006
Publication title -
journal of cellular biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.028
H-Index - 165
eISSN - 1097-4644
pISSN - 0730-2312
DOI - 10.1002/jcb.20739
Subject(s) - ap 1 transcription factor , enos , microbiology and biotechnology , biology , decoy , transcription (linguistics) , transcription factor , mutant , nitric oxide , genetics , nitric oxide synthase , endocrinology , gene , receptor , linguistics , philosophy
Abstract Human endothelial nitric oxide synthase (eNOS) plays a crucial role in maintaining blood pressure homeostasis and vascular integrity. It, therefore, is very essential to elucidate the regulation of it. In the current study, a red fluorescent protein (RFP) reporter system containing human eNOS promoter was first constructed, being characteristics of real time morphologic and quantitative analysis for the same sample. It was observed by DNA sequence deletion that 68% of the basal activity of the promoter was controlled by the region from −1 to −166 bp, and 32% of it was dependent on the region from −1,033 to −1,600 bp. The mutation of SSRE element (−999∼−994 bp) and wild‐type SSRE decoy oligodeoxynucleotides (ODN) did not alter the basal activity and the stimulating activity by lysophosphatidycholine (LPC). The mutation of upstream AP1 element (−1,530∼−1,524 bp) did not affect the basal activity, but resulted in near 30% reduction in the stimulating activity by LPC. Moreover, wild‐type AP1 decoy ODN also remarkably attenuated it. It was proved by EMSA analysis that LPC indeed enhanced the activity of AP1 transcriptional factor binding to AP1 element. However, the role of AP1 was dependent on the presence of SP1, which was proved by the combining mutation of AP1 with SP1. The mutation of downstream AP1 element (−662∼−656 bp) had no influence on the basal and stimulating activities by LPC. These results strongly suggest that the main functional region of the promoter is from −1 bp to −166 bp, that the upstream AP1 participates in the activation of the promoter by LPC on the premise of the presence of SP1, and that the downstream AP1 and SSRE do not involve the basal and stimulating activity by LPC. J. Cell. Biochem. 98: 872–884, 2006. © 2006 Wiley‐Liss, Inc.