A cAMP-responsive element binding site is essential for sterol regulation of the human lanosterol 14α-demethylase gene (CYP51)

Sunil K. Halder, Martina Fink, Michael R. Waterman, Damjana Rozman

Research output: Contribution to journalArticle

43 Citations (Scopus)

Abstract

Lanosterol 14α-demethylase (CYP51) is involved in the cholesterol biosynthesis pathway, producing follicular fluid meiosis-activating sterol. The promoter region of the human CYP51 gene contains a cluster of regulatory elements including GC box, cAMP response element (CRE), and sterol regulatory element (SRE). To understand the mechanism of sterol-dependent regulation of this gene, several constructs of the promoter with the reporter gene have been tested in JEG-3 cells containing overexpressed human sterol regulatory element binding protein (SREBP)-1a. The wild-type construct showed maximal SREBP-dependent activation, most of which is retained when the GC box is mutated/deleted. Activation is abolished when either CRE or SRE are removed/mutated. Furthermore, mutation of CRE abolishes SREBP-dependent activation after overexpression of SREBP-1a and CRE binding protein (CREB). This shows that CRE is essential, and that under ex vivo conditions CREB and SREBP cooperate in transactivating CYP51. Interestingly, protein kinase A shows a marked stimulation of the CYP51 promoter activity when overexpressed together with SREBP-1a but not when overexpressed with CREB, suggesting phosphorylation of SREBP-1a. Using a DNA probe containing all three regulatory elements, it is found that SREBP-1a, a CREB-like factor, and specificity protein (Sp1) all probably bind the CYP51 promoter. While SREBP-1a and the CRE-bound proteins are essential for the SREBP-dependent response, Sp1 apparently functions only to maximize sterol regulation of CYP51. To date this is the first gene in which cooperation between SREBP and a CREB/CRE modulator/activating transcription factor family transcription factor is shown to be essential and sufficient for SREBP-dependent activation.

Original languageEnglish (US)
Pages (from-to)1853-1863
Number of pages11
JournalMolecular Endocrinology
Volume16
Issue number8
DOIs
StatePublished - Aug 10 2002

Fingerprint

Sterol Regulatory Element Binding Proteins
Lanosterol
Sterol Regulatory Element Binding Protein 1
Sterols
Response Elements
Binding Sites
Genes
Cyclic AMP Response Element-Binding Protein
Carrier Proteins
Activating Transcription Factors
DNA Probes
Cyclic AMP-Dependent Protein Kinases
Reporter Genes
Genetic Promoter Regions
Proteins
Transcription Factors
Cholesterol
Phosphorylation
Mutation

ASJC Scopus subject areas

  • Molecular Biology
  • Endocrinology, Diabetes and Metabolism

Cite this

A cAMP-responsive element binding site is essential for sterol regulation of the human lanosterol 14α-demethylase gene (CYP51). / Halder, Sunil K.; Fink, Martina; Waterman, Michael R.; Rozman, Damjana.

In: Molecular Endocrinology, Vol. 16, No. 8, 10.08.2002, p. 1853-1863.

Research output: Contribution to journalArticle

Halder, Sunil K. ; Fink, Martina ; Waterman, Michael R. ; Rozman, Damjana. / A cAMP-responsive element binding site is essential for sterol regulation of the human lanosterol 14α-demethylase gene (CYP51). In: Molecular Endocrinology. 2002 ; Vol. 16, No. 8. pp. 1853-1863.
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abstract = "Lanosterol 14α-demethylase (CYP51) is involved in the cholesterol biosynthesis pathway, producing follicular fluid meiosis-activating sterol. The promoter region of the human CYP51 gene contains a cluster of regulatory elements including GC box, cAMP response element (CRE), and sterol regulatory element (SRE). To understand the mechanism of sterol-dependent regulation of this gene, several constructs of the promoter with the reporter gene have been tested in JEG-3 cells containing overexpressed human sterol regulatory element binding protein (SREBP)-1a. The wild-type construct showed maximal SREBP-dependent activation, most of which is retained when the GC box is mutated/deleted. Activation is abolished when either CRE or SRE are removed/mutated. Furthermore, mutation of CRE abolishes SREBP-dependent activation after overexpression of SREBP-1a and CRE binding protein (CREB). This shows that CRE is essential, and that under ex vivo conditions CREB and SREBP cooperate in transactivating CYP51. Interestingly, protein kinase A shows a marked stimulation of the CYP51 promoter activity when overexpressed together with SREBP-1a but not when overexpressed with CREB, suggesting phosphorylation of SREBP-1a. Using a DNA probe containing all three regulatory elements, it is found that SREBP-1a, a CREB-like factor, and specificity protein (Sp1) all probably bind the CYP51 promoter. While SREBP-1a and the CRE-bound proteins are essential for the SREBP-dependent response, Sp1 apparently functions only to maximize sterol regulation of CYP51. To date this is the first gene in which cooperation between SREBP and a CREB/CRE modulator/activating transcription factor family transcription factor is shown to be essential and sufficient for SREBP-dependent activation.",
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