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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

Xeroderma pigmentosum (XPF)

Jennifer Lagasca

March 4, 2003


What is xp
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


Symptoms
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)


Uv damage and ner
UV damage and NER

(www.rndsystems.com)


Review of ner
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)



Xpf ercc1 complex
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


Regions of Similarity Between XPF and ERCC1

(Gaillard et al. Nucleic Acids Research.2001)


Isolation of human xpf with rad1
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)


Xpf and ercc1 are dependent of one another
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)


R788 w mutation caucasian
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


Mutations japanese cases
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


Mutation spectrum of xpf
Mutation spectrum of XPF

(Matsumura et al. Human Molecular Genetics. 1998)


Take home note
Take home note

http://tango01.cit.nih.gov/sig/dna/drawhatis.html)