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Isolating Brown Adipocytes from Murine Interscapular Brown Adipose Tissue for Gene and Protein Expression Analysis
Published on: March 12, 2021
Human NPC1L1 expression is positively regulated by PPARα
Yuki Iwayanagi1, Tappei Takada, Fumiya Tomura
1Department of Pharmacy, The University of Tokyo Hospital Faculty of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo, 113-8655, Japan.
Pharmaceutical Research
|October 19, 2010
Summary
Peroxisome proliferator-activated receptor alpha (PPARα) directly regulates human Niemann-Pick C1-like 1 (NPC1L1) gene transcription. PPARα and PGC1α co-transactivation offers insights into glucose, fatty acid, and cholesterol metabolism.
Area of Science:
- Molecular Biology
- Metabolic Regulation
- Gene Transcription
Background:
- Niemann-Pick C1-like 1 (NPC1L1) is crucial for cholesterol absorption in enterocytes and hepatocytes.
- NPC1L1 is a key pharmacological target for cholesterol-lowering drugs like ezetimibe.
- The transcriptional regulation of NPC1L1 is not fully understood.
Purpose of the Study:
- To investigate the role of peroxisome proliferator-activated receptor alpha (PPARα) in the transcriptional regulation of human NPC1L1.
- To determine the involvement of PPARα coactivator 1α (PGC1α) in NPC1L1 gene expression.
- To identify potential regulatory elements within the human NPC1L1 gene promoter.
Main Methods:
- Reporter gene assays were used to assess promoter activity.
- Electrophoretic mobility shift assays (EMSAs) confirmed direct binding of transcription factors.
- siRNA-mediated knockdown of PPARα was employed to evaluate its effect on NPC1L1 expression.
- Deletion and mutation analyses of the NPC1L1 promoter were performed.
Main Results:
- PPARα significantly transactivated the human NPC1L1 promoter.
- A functional PPARα-response element (PPRE) was identified in the -846/-834 region of the NPC1L1 gene.
- Direct binding of PPARα and RXRα to the PPRE was confirmed by EMSA.
- Knockdown of PPARα led to decreased NPC1L1 mRNA and protein levels in HepG2 cells.
- PGC1α cotransfection enhanced NPC1L1 promoter activation mediated by SREBP2/HNF4α and PPARα/RXRα.
Conclusions:
- PPARα positively regulates human NPC1L1 transcription through direct binding to a PPRE.
- PGC1α coactivates the transcriptional regulation of human NPC1L1 by both SREBP2/HNF4α and PPARα/RXRα.
- These findings elucidate a link between PPARα, PGC1α, and NPC1L1, contributing to understanding glucose, fatty acid, and cholesterol homeostasis.
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