Characterization of three XPG-defective patients identifies three missense mutations that impair repair and...

Area of Science:

• Molecular Biology
• Genetics
• Biochemistry

Background:

• Xeroderma pigmentosum (XP) and Cockayne syndrome (CS) are rare genetic disorders.
• Mutations in the XPG gene are known to cause XP and/or CS.
• The functional impact of different XPG mutations on DNA repair and transcription is not fully understood.

Purpose of the Study:

• To investigate the functional consequences of novel XPG mutations identified in XP patients.
• To correlate specific XPG mutations with disease severity and molecular mechanisms.
• To elucidate the role of XPG in DNA repair and transcription.

Main Methods:

• Cell culture and DNA repair assays (host cell reactivation).
• XPG mRNA expression analysis.
• Genetic analysis of patient cell lines (XP40GO, XP72MA, XP165MA) and mutation characterization (p.G805R, p.W814S, p.E727X, p.L778P, p.Q150X).
• Allele-specific complementation analysis.
• Co-immunoprecipitation assays to assess protein interactions.
• Immunofluorescence to study protein recruitment to DNA damage sites.

Main Results:

• Fibroblasts from XP patients showed reduced post-UVC cell survival and impaired DNA repair, which was complemented by XPG cDNA.
• Three XPG mutations (p.G805R, p.W814S, p.L778P) were identified in the I-region of XPG.
• Missense mutations p.L778P and p.W814S retained some residual repair activity but failed to interact with TFIIH subunits (XPD, cdk7).
• Mutations destabilized XPG protein recruitment to DNA damage sites, impacting both repair and transcription.
• Severe XP/CS phenotypes were observed in patients with these novel XPG mutations.

Conclusions:

• Identified three novel XPG mutations in the I-region associated with severe XP/CS phenotypes.
• Demonstrated that these mutations impair both DNA repair and transcription.
• Highlighted the critical role of XPG's I-region in protein stability, DNA repair, and transcription regulation.