The hypocholesterolemic agent LY295427 up-regulates INSIG-1, identifying the INSIG-1 protein as a mediator of cholesterol homeostasis through SREBP.
Oxysterols regulate cholesterol homeostasis through liver X receptor (LXR; cholesterol-lowering)- and sterol regulatory element-binding protein (SREBP; cholesterol-raising)-mediated signaling pathways. Previously we reported that the hypocholesterolemic agent LY295427 (4alpha-allylcholestan-3alpha-ol) reverses oxysterol-mediated suppression of SREBP processing. We now report that LY295427 increases expression of insulin-induced gene-1 (INSIG-1) and restores SREBP processing in cells treated with oxysterols. In cells overexpressing the INSIG-1 gene, by contrast, SREBP processing is suppressed and oxysterol regulation is disrupted. SREBP processing is not restored by addition of LY295427, but is restored by increasing the levels of SREBP cleavage-activating protein (SCAP). These findings suggest that the INSIG-1 protein alters sterol balance by modulating SREBP processing jointly with SCAP. To test whether the action of oxysterols on SREBP processing is mediated through endogenous INSIG-1 protein, we used RNAi to lower the expression of the INSIG-1 gene, and found that reduced INSIG-1 protein levels caused the loss of SREBP regulation by oxysterols. We conclude that: (i) INSIG-1 gene expression is suppressed by oxysterols; (ii) LY295427 treatment counters the suppressive effects of oxysterols on SREBP processing, resulting in the expression of the INSIG-1 gene; and (iii) INSIG-1 gene expression affects SREBP processing. Taken together, these data suggest that INSIG-1 plays a critical role in regulating cholesterol concentrations in the cell.
Janowski
Bethany A
BA
Department of Molecular Genetics, University of Texas Southwestern Medical Center, Dallas, TX 75390-9046, USA. bethjanowski@charter.net
D000924:Anticholesteremic Agents / Q000494:pharmacology*; D015152:Blotting, Northern; D022762:CCAAT-Enhancer-Binding Proteins / Q000378:metabolism*; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D002777:Cholestanols / Q000494:pharmacology*; D002784:Cholesterol / Q000378:metabolism*; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005786:Gene Expression Regulation; D006706:Homeostasis; D006801:Humans; D015151:Immunoblotting; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D008954:Models, Biological; D020411:Oligonucleotide Array Sequence Analysis; D011506:Proteins / Q000378:metabolism*; D012313:RNA / Q000378:metabolism; D015398:Signal Transduction; D013256:Steroids / Q000494:pharmacology; D051780:Sterol Regulatory Element Binding Protein 1; D013997:Time Factors; D014157:Transcription Factors; D014162:Transfection; D015854:Up-Regulation
D000924:Anticholesteremic Agents; D022762:CCAAT-Enhancer-Binding Proteins; D002777:Cholestanols; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; C108626:INSIG1 protein, human; D047908:Intracellular Signaling Peptides and Proteins; C107473:LY 295427; D008565:Membrane Proteins; D011506:Proteins; C497387:SREBF1 protein, human; D013256:Steroids; D051780:Sterol Regulatory Element Binding Protein 1; D014157:Transcription Factors; D012313:RNA; D002784:Cholesterol