Selective regulation of vitamin D receptor-responsive genes by TFIIH

Area of Science:

• Molecular Biology
• Genetics
• Biochemistry

Background:

• Xeroderma pigmentosum (XP) is a genetic disorder caused by mutations in DNA repair genes.
• The transcription/repair factor TFIIH, containing the XPD subunit, is crucial for DNA repair and transcription.
• Vitamin D receptor (VDR) is a nuclear receptor that regulates gene expression.

Purpose of the Study:

• To investigate the molecular mechanisms underlying VDR transactivation defects in XP-D cells.
• To determine the role of TFIIH and its subunits in VDR-mediated gene regulation.
• To elucidate the specific interactions between VDR, TFIIH, and other transcription factors.

Main Methods:

• Cell culture of XP-D deficient cells.
• Gene expression analysis of VDR target genes (e.g., CYP24).
• Chromatin immunoprecipitation (ChIP) assays to assess protein recruitment to promoters.
• In vitro kinase assays to study phosphorylation of TFIIH components and activators.

Main Results:

• XP-D cells exhibit selective inhibition of VDR transactivation for specific genes like CYP24.
• The XPD/R683W mutation impairs VDR recruitment to the CYP24 promoter.
• Unlike other nuclear receptors, VDR is not phosphorylated by TFIIH's cdk7 kinase.
• The defect in XP-D cells is attributed to impaired phosphorylation of the activator Ets1 by TFIIH.
• Phosphorylated Ets1 is essential for VDR binding and subsequent recruitment of coactivators and RNA polymerase II.

Conclusions:

• TFIIH regulates VDR activity through phosphorylation of either VDR's A/B domain or a DNA-binding partner like Ets1.
• The study reveals a novel mechanism of nuclear receptor regulation involving TFIIH-mediated phosphorylation of accessory factors.
• This finding provides insights into the pathogenesis of XP-D and the intricate regulation of gene expression by nuclear receptors.