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Analysis of The HMG‐CoA Reductase Promoter by in vivo Electroporation
Author(s) -
Lagor William R.,
Heller Richard,
Ness Gene C.
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.20.5.lb43-b
Subject(s) - luciferase , reductase , electroporation , in vivo , hmg coa reductase , biology , chemistry , microbiology and biotechnology , biochemistry , transfection , enzyme , gene , genetics
HMG‐CoA reductase catalyzes the rate‐limiting step in cholesterol biosynthesis. Its expression is under strict regulation by a number of dietary and hormonal stimuli. Previous work on this promoter has been performed primarily in vitro in a number of different tumor or transformed cells. However the liver is the primary site of regulatable cholesterol synthesis in the body and thus is responsible for cholesterol balance. Therefore it is of paramount importance to study regulation of HMG‐CoA reductase in liver. Do to this we have used in vivo electroporation to deliver HMG‐CoA reductase promoter‐luciferase reporter constructs directly into the livers of rats. A series of 6 sequential deletion constructs were introduced at different sites in the liver of the same animal for direct comparison. Luciferase activity was markedly and significantly increased with the −123 construct compared to the −58 construct. This region contains CRE and NF‐Y sites previously shown by in vivo footprinting to bind transcription factors. These sites may be important for regulation by insulin. A significant increase in luciferase activity was also observed with the −325 construct as compared to the −228 construct. A LRH‐1/FTF site that may be involved in regulation by bile acids lies in this region. The data show that in vivo electroporation is a viable and useful method for studying promoter function in live animals. (Supported by grant 04TSP‐03 from the Florida Department of Health.)