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Xeroderma pigmentosum (XPF)

Xeroderma pigmentosum (XPF). Jennifer Lagasca March 4, 2003. What is XP?. Greek for “dry skin” (first case in 1870) inability to remove UV-induced damage from their DNA (1000 fold increased risk of cancer) autosomal recessive 16p13.1-13.2

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Xeroderma pigmentosum (XPF)

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  1. Xeroderma pigmentosum (XPF) Jennifer Lagasca March 4, 2003

  2. What is XP? • Greek for “dry skin” (first case in 1870) • inability to remove UV-induced damage from their DNA (1000 fold increased risk of cancer) • autosomal recessive • 16p13.1-13.2 • 7 complementation groups XPA-XPG (range in prevalence/severity) • prevalence (1:250000 US/Europe; 1:40000 Japan) • doesn’t discriminate

  3. Symptoms • 1. Pigmentary changes (reddened/scale skin); irregular dark spots • 2. Poikiloderma-irregular patches of light and dark spots (spiderweb/thinning) • 3. Premature skin aging, dev. Of solar keratoses and skin cancers • other related symptoms: eye problems (lesions) (80%); neurological problems (20%) • XPF-mild phenotype USUALLY w/o neurological abnormalities • treatment (Steeg et al. Molecular Medicine Today.1999)

  4. UV damage and NER (www.rndsystems.com)

  5. Review of NER • DNA damage recognition: XPC/HHR23B, XPA, XPE • DNA helix opening: XPB and XPD (helicases) • Dual incision: XPG (3’end) and XPF/ERCC1 (5’end) • Excision: DNA polymerase removes excised oligonucleotide • Gapfilling: DNA ligase fills gap by DNA synthesis • (www.rndsystems.com)

  6. (Steeg et al.Molecular Medicine Today.1999)

  7. XPF/ERCC1 complex (Gaillard et al. Nucleic Acids Research. 2001) • XPF (103kDa) and ERCC1 (31kDa) incise at 22 nt to 5’ • side of lesion • -N-terminal (DNA binding and endonuclease activity) • C-terminal (binds to ERCC1)-significant sequence similarity

  8. Regions of Similarity Between XPF and ERCC1 (Gaillard et al. Nucleic Acids Research.2001)

  9. Isolation of human XPF with Rad1 • S.cerevisiae (RAD1/RAD10 complex) -> cleave 5’ end • used primers (homology sequence) residues 699-758 (RT-PCR) - as probe • localized to 16p13.1-13.2 • verified by complementation test -> injected isolated gene in nuclei of XPF pt.’s cells • cells irradiated with UV damage and radioactive letters • increased number of grains above the nuclei-> restored repair activity to normal level (Sijbers et al. Cell.1996)

  10. XPF and ERCC1 are dependent of one another (Yagi et al. Carcinogenesis.1998) • *anti-ERCC1 ab coimmunoprecipitated • a 120 kDa protein from normal cell • extract but not XPF cell extract (1997) • *low content of ERCC1 and 120 kDA • protein (frequent feature of XPF patients) • *transfected XP-F cDNA into mammalian vector plasmid -> XPFcells(1998) • *clone XP-FR2 -expressed high levels of ERCC1 and XPF (shows that ERCC1 is stabilized by binding to XPF)

  11. R788 W Mutation (Caucasian) • 2nd Caucasian XPF patient (XP42RO)-late 40s w/ neurological symptoms (4-6 times reductions of NER activity) • Homozygous for point mutation in XPF (C->T at nuc.2377 changes arginine 788 to tryptophan • This mutation is leaky because a small amount of XPF could still be detected – still has repair activity-explains the mild phenotype • This same mutation was found in another Caucasian XP-F case; perhaps a Caucasian founder mutation? Sijbers et al. The Journal of Investigative Dermatology.1998

  12. Mutations (Japanese cases) • 3 types of mutations: 1) aa subs, 2) aa/truncated, 3) truncated • N-terminal (homologous 86-333)-(XP101OS-heterozygous pt. mutation A-> G); all XPF cells have at least 1 normal allele in this region-suggests this is essential for basic cellular functioning • C-terminal (homologous 559-823)-ERCC1 binding domain • Mutations (XP3YO,XP2YO,XP1TS) • XP23OS lack this region- but still have XPF phenotype (mutation in C-terminal does not end NER function; supports that N-terminal (1-443) is important for process of NER) • XP24KY(truncated protein (no C-terminal) and protein with single aa subs (R443W) • Truncation probably not responsible for neurological disorders (XP23OS);R443W might • **mutations of XPF affecting C-terminal region-interfere with complex formation->rapid degradation of ERCC1

  13. Mutation spectrum of XPF (Matsumura et al. Human Molecular Genetics. 1998)

  14. Take home note http://tango01.cit.nih.gov/sig/dna/drawhatis.html)

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