Gene Summary

Gene:POLH; polymerase (DNA directed), eta
Aliases: XPV, XP-V, RAD30, RAD30A
Summary:This gene encodes a member of the Y family of specialized DNA polymerases. It copies undamaged DNA with a lower fidelity than other DNA-directed polymerases. However, it accurately replicates UV-damaged DNA; when thymine dimers are present, this polymerase inserts the complementary nucleotides in the newly synthesized DNA, thereby bypassing the lesion and suppressing the mutagenic effect of UV-induced DNA damage. This polymerase is thought to be involved in hypermutation during immunoglobulin class switch recombination. Mutations in this gene result in XPV, a variant type of xeroderma pigmentosum. Several transcript variants encoding different isoforms have been found for this gene. [provided by RefSeq, May 2014]
Databases:OMIM, VEGA, HGNC, Ensembl, GeneCard, Gene
Protein:DNA polymerase eta
Source:NCBIAccessed: 11 August, 2015


What does this gene/protein do?
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Pathways:What pathways are this gene/protein implicaed in?
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Cancer Overview

Research Indicators

Publications Per Year (1990-2015)
Graph generated 11 August 2015 using data from PubMed using criteria.

Literature Analysis

Mouse over the terms for more detail; many indicate links which you can click for dedicated pages about the topic.

  • Codon, Nonsense
  • Ultraviolet Rays
  • Adolescents
  • Molecular Sequence Data
  • DNA-Binding Proteins
  • Childhood Cancer
  • Exons
  • Genetic Variation
  • Squamous Cell Carcinoma
  • Apoptosis
  • Mutation
  • Skin Cancer
  • DNA-Directed DNA Polymerase
  • Fibroblasts
  • DNA Damage
  • Pedigree
  • Genotype
  • DNA Replication
  • Pyrimidine Dimers
  • Missense Mutation
  • Models, Molecular
  • Western Blotting
  • Chromosome 6
  • Gene Expression
  • Tumor Markers
  • siRNA
  • Caffeine
  • Cell Cycle Proteins
  • Cultured Cells
  • Xeroderma Pigmentosum Group D Protein
  • Messenger RNA
  • Base Sequence
  • Genomics
  • Protein Structure, Tertiary
  • Case-Control Studies
  • Nucleic Acid Synthesis Inhibitors
  • DNA
  • DNA Repair
  • European Continental Ancestry Group
Tag cloud generated 11 August, 2015 using data from PubMed, MeSH and CancerIndex

Specific Cancers (1)

Data table showing topics related to specific cancers and associated disorders. Scope includes mutations and abnormal protein expression.

Note: list is not exhaustive. Number of papers are based on searches of PubMed (click on topic title for arbitrary criteria used).

Latest Publications: POLH (cancer-related)

Williams LN, Marjavaara L, Knowels GM, et al.
dNTP pool levels modulate mutator phenotypes of error-prone DNA polymerase ε variants.
Proc Natl Acad Sci U S A. 2015; 112(19):E2457-66 [PubMed] Article available free on PMC after 12/11/2015 Related Publications
Mutator phenotypes create genetic diversity that fuels tumor evolution. DNA polymerase (Pol) ε mediates leading strand DNA replication. Proofreading defects in this enzyme drive a number of human malignancies. Here, using budding yeast, we show that mutator variants of Pol ε depend on damage uninducible (Dun)1, an S-phase checkpoint kinase that maintains dNTP levels during a normal cell cycle and up-regulates dNTP synthesis upon checkpoint activation. Deletion of DUN1 (dun1Δ) suppresses the mutator phenotype of pol2-4 (encoding Pol ε proofreading deficiency) and is synthetically lethal with pol2-M644G (encoding altered Pol ε base selectivity). Although pol2-4 cells cycle normally, pol2-M644G cells progress slowly through S-phase. The pol2-M644G cells tolerate deletions of mediator of the replication checkpoint (MRC) 1 (mrc1Δ) and radiation sensitive (Rad) 9 (rad9Δ), which encode mediators of checkpoint responses to replication stress and DNA damage, respectively. The pol2-M644G mutator phenotype is partially suppressed by mrc1Δ but not rad9Δ; neither deletion suppresses the pol2-4 mutator phenotype. Thus, checkpoint activation augments the Dun1 effect on replication fidelity but is not required for it. Deletions of genes encoding key Dun1 targets that negatively regulate dNTP synthesis, suppress the dun1Δ pol2-M644G synthetic lethality and restore the mutator phenotype of pol2-4 in dun1Δ cells. DUN1 pol2-M644G cells have constitutively high dNTP levels, consistent with checkpoint activation. In contrast, pol2-4 and POL2 cells have similar dNTP levels, which decline in the absence of Dun1 and rise in the absence of the negative regulators of dNTP synthesis. Thus, dNTP pool levels correlate with Pol ε mutator severity, suggesting that treatments targeting dNTP pools could modulate mutator phenotypes for therapy.

Japp AS, Gessi M, Messing-Jünger M, et al.
High-resolution genomic analysis does not qualify atypical plexus papilloma as a separate entity among choroid plexus tumors.
J Neuropathol Exp Neurol. 2015; 74(2):110-20 [PubMed] Related Publications
Choroid plexus tumors are rare neoplasms that mainly affect children. They include papillomas, atypical papillomas, and carcinomas. Detailed genetic studies are rare, and information about their molecular pathogenesis is limited. Molecular inversion probe analysis is a hybridization-based method that represents a reliable tool for the analysis of highly fragmented formalin-fixed paraffin-embedded tissue-derived DNA. Here, analysis of 62 cases showed frequent hyperdiploidy in papillomas and atypical papillomas that appeared very similar in their cytogenetic profiles. In contrast, carcinomas showed mainly losses of chromosomes. Besides recurrent focal chromosomal gains common to all choroid plexus tumors, including chromosome 14q21-q22 (harboring OTX2), chromosome 7q22 (LAMB1), and chromosome 9q21.12 (TRPM3), Genomic Identification of Significant Targets in Cancer analysis uncovered focal alterations specific for papillomas and atypical papillomas (e.g. 7p21.3 [ARL4A]) and for carcinomas (16p13.3 [RBFOX1] and 6p21 [POLH, GTPBP2, RSPH9, and VEGFA]). Additional RNA expression profiling and gene set enrichment analysis revealed greater expression of cell cycle-related genes in atypical papillomas in comparison with that in papillomas. These findings suggest that atypical papillomas represent an immature variant of papillomas characterized by increased proliferative activity, whereas carcinomas seem to represent a genetically distinct tumor group.

Ben Rekaya M, Laroussi N, Messaoud O, et al.
A founder large deletion mutation in Xeroderma pigmentosum-Variant form in Tunisia: implication for molecular diagnosis and therapy.
Biomed Res Int. 2014; 2014:256245 [PubMed] Article available free on PMC after 12/11/2015 Related Publications
Xeroderma pigmentosum Variant (XP-V) form is characterized by a late onset of skin symptoms. Our aim is the clinical and genetic investigations of XP-V Tunisian patients in order to develop a simple tool for early diagnosis. We investigated 16 suspected XP patients belonging to ten consanguineous families. Analysis of the POLH gene was performed by linkage analysis, long range PCR, and sequencing. Genetic analysis showed linkage to the POLH gene with a founder haplotype in all affected patients. Long range PCR of exon 9 to exon 11 showed a 3926 bp deletion compared to control individuals. Sequence analysis demonstrates that this deletion has occurred between two Alu-Sq2 repetitive sequences in the same orientation, respectively, in introns 9 and 10. We suggest that this mutation POLH NG_009252.1: g.36847_40771del3925 is caused by an equal crossover event that occurred between two homologous chromosomes at meiosis. These results allowed us to develop a simple test based on a simple PCR in order to screen suspected XP-V patients. In Tunisia, the prevalence of XP-V group seems to be underestimated and clinical diagnosis is usually later. Cascade screening of this founder mutation by PCR in regions with high frequency of XP provides a rapid and cost-effective tool for early diagnosis of XP-V in Tunisia and North Africa.

Herman KN, Toffton S, McCulloch SD
Detrimental effects of UV-B radiation in a xeroderma pigmentosum-variant cell line.
Environ Mol Mutagen. 2014; 55(5):375-84 [PubMed] Article available free on PMC after 12/11/2015 Related Publications
DNA polymerase η (pol η), of the Y-family, is well known for its in vitro DNA lesion bypass ability. The most well-characterized lesion bypassed by this polymerase is the cyclobutane pyrimidine dimer (CPD) caused by ultraviolet (UV) light. Historically, cellular and whole-animal models for this area of research have been conducted using UV-C (λ=100-280 nm) owing to its ability to generate large quantities of CPDs and also the more structurally distorting 6-4 photoproduct. Although UV-C is useful as a laboratory tool, exposure to these wavelengths is generally very low owing to being filtered by stratospheric ozone. We are interested in the more environmentally relevant wavelength range of UV-B (λ=280-315 nm) for its role in causing cytotoxicity and mutagenesis. We evaluated these endpoints in both a normal human fibroblast control line and a Xeroderma pigmentosum variant cell line in which the POLH gene contains a truncating point mutation, leading to a nonfunctional polymerase. We demonstrate that UV-B has similar but less striking effects compared to UV-C in both its cytotoxic and its mutagenic effects. Analysis of the mutation spectra after a single dose of UV-B shows that a majority of mutations can be attributed to mutagenic bypass of dipyrimidine sequences. However, we do note additional types of mutations with UV-B that are not previously reported after UV-C exposure. We speculate that these differences are attributed to a change in the spectra of photoproduct lesions rather than other lesions caused by oxidative stress.

Buisson R, Niraj J, Pauty J, et al.
Breast cancer proteins PALB2 and BRCA2 stimulate polymerase η in recombination-associated DNA synthesis at blocked replication forks.
Cell Rep. 2014; 6(3):553-64 [PubMed] Article available free on PMC after 12/11/2015 Related Publications
One envisioned function of homologous recombination (HR) is to find a template for DNA synthesis from the resected 3'-OH molecules that occur during double-strand break (DSB) repair at collapsed replication forks. However, the interplay between DNA synthesis and HR remains poorly understood in higher eukaryotic cells. Here, we reveal functions for the breast cancer proteins BRCA2 and PALB2 at blocked replication forks and show a role for these proteins in stimulating polymerase η (Polη) to initiate DNA synthesis. PALB2, BRCA2, and Polη colocalize at stalled or collapsed replication forks after hydroxyurea treatment. Moreover, PALB2 and BRCA2 interact with Polη and are required to sustain the recruitment of Polη at blocked replication forks. PALB2 and BRCA2 stimulate Polη-dependent DNA synthesis on D loop substrates. We conclude that PALB2 and BRCA2, in addition to their functions in D loop formation, play crucial roles in the initiation of recombination-associated DNA synthesis by Polη-mediated DNA repair.

Zhou W, Chen YW, Liu X, et al.
Expression of DNA translesion synthesis polymerase η in head and neck squamous cell cancer predicts resistance to gemcitabine and cisplatin-based chemotherapy.
PLoS One. 2013; 8(12):e83978 [PubMed] Article available free on PMC after 12/11/2015 Related Publications
PURPOSE: The development of resistance against anticancer drugs has been a persistent clinical problem for the treatment of locally advanced malignancies in the head and neck mucosal derived squamous cell carcinoma (HNSCC). Recent evidence indicates that the DNA translesion synthesis (TLS) polymerase η (Pol η; hRad30a gene) reduces the effectiveness of gemcitabine/cisplatin. The goal of this study is to examine the relationship between the expression level of Pol η and the observed resistance against these chemotherapeutic agents in HNSCC, which is currently unknown.
METHODS: Sixty-four mucosal derived squamous cell carcinomas of head and neck (HNSCC) from 1989 and 2007 at the City of Hope National Medical Center (Duarte, CA) were retrospectively analyzed. Pretreatment samples were immunostained with anti-Pol η antibody and the correlation between the expression level of Pol η and clinical outcomes were evaluated. Forty-nine cases treated with platinum (n=40) or gemcitabine (n=9) based chemotherapy were further examined for Pol η expression level for comparison with patient response to chemotherapy.
RESULTS: The expression of Pol η was elevated in 67% of the head and neck tumor samples. Pol η expression level was significantly higher in grade 1 to grade 2 tumors (well to moderately differentiated). The overall benefit rate (complete response+ partial response) in patients treated with platinum and gemcitabine based chemotherapy was 79.5%, where low Pol η level was significantly associated with high complete response rate (p=0.03), although not associated with overall survival. Furthermore, no significant correlation was observed between Pol η expression level with gender, age, tobacco/alcohol history, tumor stage and metastatic status.
CONCLUSIONS: Our data suggest that Pol η expression may be a useful prediction marker for the effectiveness of platinum or gemcitabine based therapy for HNSCC.

Opletalova K, Bourillon A, Yang W, et al.
Correlation of phenotype/genotype in a cohort of 23 xeroderma pigmentosum-variant patients reveals 12 new disease-causing POLH mutations.
Hum Mutat. 2014; 35(1):117-28 [PubMed] Related Publications
Xeroderma pigmentosum variant (XP-V) is a rare genetic disease, characterized by some sunlight sensitivity and predisposition to cutaneous malignancies. We described clinical and genetic features of the largest collection ever published of 23 XPV patients (ages between 21 and 86) from 20 unrelated families. Primary fibroblasts from patients showed normal nucleotide excision repair but UV-hypersensitivity in the presence of caffeine, a signature of the XP-V syndrome. 87% of patients developed skin tumors with a median age of 21 for the first occurrence. The median numbers of basal-cell carcinoma was 13 per patient, six for squamous-cell carcinoma, and five for melanoma. XP-V is due to defects in the translesion-synthesis DNA polymerase Polη coded by the POLH gene. DNA sequencing of POLH revealed 29 mutations, where 12 have not been previously identified, leading to truncated polymerases in 69% of patients. Four missense mutations are correlated with the protein stability by structural modeling of the Polη polymerase domain. There is a clear relationship between the types of missense mutations and clinical severity. For truncating mutations, which lead to an absence of or to inactive proteins, the life-cumulated UV exposure is probably the best predictor of cancer incidence, reinforcing the necessity to protect XP-Vs from sun exposure.

Ortega-Recalde O, Vergara JI, Fonseca DJ, et al.
Whole-exome sequencing enables rapid determination of xeroderma pigmentosum molecular etiology.
PLoS One. 2014; 8(6):e64692 [PubMed] Article available free on PMC after 12/11/2015 Related Publications
Xeroderma pigmentosum (XP) is a rare autosomal recessive disorder characterized by extreme sensitivity to actinic pigmentation changes in the skin and increased incidence of skin cancer. In some cases, patients are affected by neurological alterations. XP is caused by mutations in 8 distinct genes (XPA through XPG and XPV). The XP-V (variant) subtype of the disease results from mutations in a gene (XPV, also named POLH) which encodes for Polη, a member of the Y-DNA polymerase family. Although the presence and severity of skin and neurological dysfunctions differ between XP subtypes, there are overlapping clinical features among subtypes such that the sub-type cannot be deduced from the clinical features. In this study, in order to overcome this drawback, we undertook whole-exome sequencing in two XP sibs and their father. We identified a novel homozygous nonsense mutation (c.897T>G, p.Y299X) in POLH which causes the disease. Our results demonstrate that next generation sequencing is a powerful approach to rapid determination of XP genetic etiology.

Ono R, Masaki T, Takeuchi S, et al.
Three school-age cases of xeroderma pigmentosum variant type.
Photodermatol Photoimmunol Photomed. 2013; 29(3):132-9 [PubMed] Related Publications
BACKGROUND: Xeroderma pigmentosum (XP) is a photosensitive genodermatosis with increased susceptibility to skin cancers. Patients are typically diagnosed with XP when they consult a dermatologist for skin cancers.
CASE/METHODS: The genetic analysis and 2-8 years of follow-up for three school-age patients with XP-V is described. The patients were referred to us because of increased pigmented freckles; they had not experienced abnormal sunburn or developed skin cancer at their first visit. All patients harbored a genetic mutation in the POLH gene. XPV9KO was diagnosed at age 13 with a homozygous del1661A that creates a stop codon in the non-catalytic domain of POLH. The patient practiced sun protection, effectively preventing the development of skin cancer by age 21. XPV19KO was diagnosed at age 11 with a compound heterozygous mutation of G490T and C1066T, causing POLH truncation in the catalytic domain. This patient developed basal cell carcinoma at ages 12 and 13. XPV18KO was referred to us at age 11 and diagnosed with compound heterozygous variants of c.1246_1311del66 (exon 9 skipping), a novel mutation, and c.661_764 del104 (exon 6 skipping).
CONCLUSION: Freckle-like pigmentation on sun-exposed skin is sometimes the only sign of XP-V, and early diagnosis is extremely important for children.

Muller HK, Woods GM
Ultraviolet radiation effects on the proteome of skin cells.
Adv Exp Med Biol. 2013; 990:111-9 [PubMed] Related Publications
Proteomic studies to date have had limited use as an investigative tool in the skin's response to UV radiation. These studies used cell lines and reconstructed skin and have shown evidence of cell injury with oxidative damage and stress induced heat shock proteins. Others changes included altered cytokeratin and cytoskeletal proteins with enhanced expression of TRIM29 as the keratinocytes regenerate. The associated DNA repair requires polη, Rad18/Rad16 and Rev1. In the whole animal these events would be associated with inflammation, remodelling of the epidermis and modulation of the immune response. Longer term changes include ageing and skin cancers such as melanoma, squamous cell carcinoma and basal cell carcinoma. In the future proteomics will be used to explore these important aspects of photobiology. Better characterisation of the proteins involved should lead to a greater understanding of the skin's response to UV radiation.

Durando M, Tateishi S, Vaziri C
A non-catalytic role of DNA polymerase η in recruiting Rad18 and promoting PCNA monoubiquitination at stalled replication forks.
Nucleic Acids Res. 2013; 41(5):3079-93 [PubMed] Article available free on PMC after 12/11/2015 Related Publications
Trans-lesion DNA synthesis (TLS) is a DNA damage-tolerance mechanism that uses low-fidelity DNA polymerases to replicate damaged DNA. The inherited cancer-propensity syndrome xeroderma pigmentosum variant (XPV) results from error-prone TLS of UV-damaged DNA. TLS is initiated when the Rad6/Rad18 complex monoubiquitinates proliferating cell nuclear antigen (PCNA), but the basis for recruitment of Rad18 to PCNA is not completely understood. Here, we show that Rad18 is targeted to PCNA by DNA polymerase eta (Polη), the XPV gene product that is mutated in XPV patients. The C-terminal domain of Polη binds to both Rad18 and PCNA and promotes PCNA monoubiquitination, a function unique to Polη among Y-family TLS polymerases and dissociable from its catalytic activity. Importantly, XPV cells expressing full-length catalytically-inactive Polη exhibit increased recruitment of other error-prone TLS polymerases (Polκ and Polι) after UV irradiation. These results define a novel non-catalytic role for Polη in promoting PCNA monoubiquitination and provide a new potential mechanism for mutagenesis and genome instability in XPV individuals.

Senhaji MA, Abidi O, Nadifi S, et al.
c.1643_1644delTG XPC mutation is more frequent in Moroccan patients with xeroderma pigmentosum.
Arch Dermatol Res. 2013; 305(1):53-7 [PubMed] Related Publications
Xeroderma pigmentosum is a rare autosomal recessive disease characterized by hypersensitivity to UV light which is due to alterations of the nucleotide excision repair pathway. Eight genes (XPA to XPG and XPV) are responsible for the disease. Among them, the XPC gene is known to be the most mutated in Mediterranean patients. The aim of this study was to determine the frequency of the most common XPC mutation and describe the clinical features of Moroccan patients with xeroderma pigmentosum. Twenty four patients belonging to 21 unrelated Moroccan families and 58 healthy subjects were investigated. After clinical examination, the screening for the c.1643_1644delTG (p.Val548AlafsX25) mutation in the XPC gene was performed by PCR and automated sequencing of exon 9 in all patients and controls. The molecular analysis showed that among the 24 patients, 17 were homozygous for the c.1643_1644delTG mutation and all their tested parents were heterozygous, whereas the others (7 patients) did not carry the mutation. The frequency of this mutation was estimated to be 76.19 % (16/21 families). None of the 58 healthy individuals carried this mutation. In addition, clinical investigation showed that the majority of the patients bearing this mutation have the same clinical features. Our results revealed that the p.Val548AlafsX25 mutation is the major cause (76.19 %) of xeroderma pigmentosum in Moroccan families. This would have an important impact on improving management of patients and their relatives.

Fassihi H
Spotlight on 'xeroderma pigmentosum'.
Photochem Photobiol Sci. 2013; 12(1):78-84 [PubMed] Related Publications
Xeroderma pigmentosum (XP) is a rare autosomal recessive disorder of DNA repair characterised by photosensitivity, progressive pigmentary change, and an increased incidence of ultraviolet (UV)-induced skin and mucous membrane cancers. Approximately 25% of XP patients also have progressive neurological degeneration. There are eight XP complementation groups (XP-A through to XP-G, and XP variant (XP-V)), corresponding to the affected DNA repair gene. Seven of these genes, XPA to XPG, are involved in nucleotide excision repair, removing UV-induced damage from DNA. The eighth gene, XPV (or POLH), encodes for DNA polymerase η, which is required for the replication of DNA containing unrepaired UV-induced damage. There is wide variability in clinical features both between and within XP complementation groups. The diagnosis is made clinically and confirmed by cellular tests for defective DNA repair. This is followed by identification of the defective gene (complementation analysis) and causative mutation(s). Although there is no cure, sun avoidance and regular follow-up to assess and treat any skin cancers increase life expectancy. The neurological abnormalities are progressive and result in a shortened lifespan. The study of patients with XP has highlighted the importance of nucleotide excision repair in the aetiology of skin cancers and neurological degeneration, and has solidified the link between UV exposure, DNA damage, somatic mutations and skin cancer.

Lehmann AR, McGibbon D, Stefanini M
Xeroderma pigmentosum.
Orphanet J Rare Dis. 2011; 6:70 [PubMed] Article available free on PMC after 12/11/2015 Related Publications
Xeroderma pigmentosum (XP) is defined by extreme sensitivity to sunlight, resulting in sunburn, pigment changes in the skin and a greatly elevated incidence of skin cancers. It is a rare autosomal recessive disorder and has been found in all continents and racial groups. Estimated incidences vary from 1 in 20, 000 in Japan to 1 in 250, 000 in the USA, and approximately 2.3 per million live births in Western Europe.The first features are either extreme sensitivity to sunlight, triggering severe sunburn, or, in patients who do not show this sun-sensitivity, abnormal lentiginosis (freckle-like pigmentation due to increased numbers of melanocytes) on sun-exposed areas. This is followed by areas of increased or decreased pigmentation, skin aging and multiple skin cancers, if the individuals are not protected from sunlight. A minority of patients show progressive neurological abnormalities. There are eight XP complementation groups, corresponding to eight genes, which, if defective, can result in XP. The products of these genes are involved in the repair of ultraviolet (UV)-induced damage in DNA. Seven of the gene products (XPA through G) are required to remove UV damage from the DNA. The eighth (XPV or DNA polymerase η) is required to replicate DNA containing unrepaired damage. There is wide variability in clinical features both between and within XP groups. Diagnosis is made clinically by the presence, from birth, of an acute and prolonged sunburn response at all exposed sites, unusually early lentiginosis in sun-exposed areas or onset of skin cancers at a young age. The clinical diagnosis is confirmed by cellular tests for defective DNA repair. These features distinguish XP from other photodermatoses such as solar urticaria and polymorphic light eruption, Cockayne Syndrome (no pigmentation changes, different repair defect) and other lentiginoses such as Peutz-Jeghers syndrome, Leopard syndrome and Carney complex (pigmentation not sun-associated), which are inherited in an autosomal dominant fashion. Antenatal diagnosis can be performed by measuring DNA repair or by mutation analysis in CVS cells or in amniocytes. Although there is no cure for XP, the skin effects can be minimised by rigorous protection from sunlight and early removal of pre-cancerous lesions. In the absence of neurological problems and with lifetime protection against sunlight, the prognosis is good. In patients with neurological problems, these are progressive, leading to disabilities and a shortened lifespan.

Hentosh P, Benjamin T, Hall L, et al.
Xeroderma pigmentosum variant: complementary molecular approaches to detect a 13 base pair deletion in the DNA polymerase eta gene.
Exp Mol Pathol. 2011; 91(2):528-33 [PubMed] Related Publications
Deficiencies of DNA polymerase eta-an enzyme mediating replication past UV-induced DNA damage-predispose individuals to xeroderma pigmentosum variant (XPV) and result in a high incidence of skin cancers. We designed, developed and assessed several complementary molecular approaches to detect a genetically inherited deletion within DNA polymerase eta. RNA was reverse transcribed from XPV fibroblasts and from normal human cells, and standard polymerase chain reaction (PCR) was conducted on the cDNA targeting a region with a 13 base pair deletion within the polymerase eta gene. PCR products were subjected to restriction fragment length polymorphism (RFLP) analysis and cycle DNA sequencing. The deletion was found to eliminate a BsrGI restriction site and affected the number of resultant fragments visualized after gel electrophoresis. Cycle sequencing of polymerase eta-specific amplicons from XPV and normal cells provided a second approach for detecting the mutation. Additionally, the use of a fluorescent nucleic acid dye-EvaGreen-in real-time PCR and melt curve analysis distinguished normal and XPV patient-derived amplicons as well as heteroduplexes that represent heterozygotic carriers without the need for high resolution melt analysis-compatible software. Our approaches are easily adaptable by diagnostic laboratories that screen for or verify genetically inherited disorders and identify carriers of a defective gene.

Välk K, Vooder T, Kolde R, et al.
Gene expression profiles of non-small cell lung cancer: survival prediction and new biomarkers.
Oncology. 2010; 79(3-4):283-92 [PubMed] Related Publications
OBJECTIVES: Despite the well-defined histological types of non-small cell lung cancer (NSCLC), a given stage is often associated with wide-ranging survival rates and treatment outcomes. This disparity has led to an increased demand for the discovery and identification of new informative biomarkers.
METHODS: In the current study, we screened 81 NSCLC samples using Illumina whole-genome gene expression microarrays in an effort to identify differentially expressed genes and new NSCLC biomarkers.
RESULTS: We identified novel genes whose expression was upregulated in NSCLC, including SPAG5, POLH, KIF23, and RAD54L, which are associated with mitotic spindle formation, DNA repair, chromosome segregation, and dsDNA break repair, respectively. We also identified several novel genes whose expression was downregulated in NSCLC, including SGCG, NLRC4, MMRN1, and SFTPD, which are involved in extracellular matrix formation, apoptosis, blood vessel leakage, and inflammation, respectively. We found a significant correlation between RNA degradation and survival in adenocarcinoma cases.
CONCLUSIONS: Even though the follow-up time was too limited to draw final conclusions, we were able to show better prediction p values in a group selection based on molecular profiles compared to histology. The current study also uncovered new candidate biomarker genes that are likely to be involved in diverse processes associated with NSCLC development.

Chou KM
DNA polymerase eta and chemotherapeutic agents.
Antioxid Redox Signal. 2011; 14(12):2521-9 [PubMed] Article available free on PMC after 12/11/2015 Related Publications
The discovery of human DNA polymerase eta (pol η) has a major impact on the fields of DNA replication/repair fields. Since the discovery of human pol η, a number of new DNA polymerases with the ability to bypass various DNA lesions have been discovered. Among these polymerases, pol η is the most extensively studied lesion bypass polymerase with a defined major biological function, that is, to replicate across the cyclobutane pyrimidine dimers introduced by UV irradiation. Cyclobutane pyrimidine dimer is a major DNA lesion that causes distortion of DNA structure and block the replicative DNA polymerases during DNA replication process. Genetic defects in the pol η gene, Rad30, results in a disease called xeroderma pigmentosum variant. This review focuses on the overall properties of pol η and the mechanism that involved in regulating its activity in cells. In addition, the role of pol η in the action of DNA-targeting anticancer compounds is also discussed.

Messaoud O, Ben Rekaya M, Cherif W, et al.
Genetic homogeneity of mutational spectrum of group-A xeroderma pigmentosum in Tunisian patients.
Int J Dermatol. 2010; 49(5):544-8 [PubMed] Related Publications
BACKGROUND: Xeroderma Pigmentosum (XP) is a rare autosomal recessive disorder characterized by cutaneous and ocular alterations. Eight genes, Xeroderma Pigmentosum group A (XPA) to Xeroderma Pigmentosum group G (XPG) and Xeroderma Pigmentosum group V (XPV), are known to be responsible for the disease and products of these genes are involved in the repair of deoxyribonucleic acid (DNA) lesions generated by UV radiation. Several XP patients suffer from neurological defects, found in the XPA (the most common form), D and G groups. The aim of this study was to investigate the mutational spectrum of XPA in Tunisia, in order to propose a simple tool for molecular diagnosis.
METHODS: This study was carried out on six unrelated families with nine Tunisian XPA patients. Clinical features were recorded. As a previous study showed the presence of the R228X mutation in Tunisia, patients were first screened for this mutation by polymerase chain reaction-restriction fragment length polymorphism and then confirmed by direct sequencing.
RESULTS: The results showed that all patients carried the XPA R228X mutation. This mutation corresponds to a C to T transition, which creates a premature stop codon at position 228, thus causing a DNA repair defect.
CONCLUSIONS: The XPA R228X mutation is common in Tunisian population. This mutation is associated with a relatively moderate phenotype of the XPA. As all explored patients presented the recurrent mutation XPA R228X, a potential founder effect was searched and confirmed by haplotype analysis. Taking into account similar genetic background, investigation of this mutation should allow a cost effective and rapid diagnosis of XPA in north-African populations.

Ziv O, Geacintov N, Nakajima S, et al.
DNA polymerase zeta cooperates with polymerases kappa and iota in translesion DNA synthesis across pyrimidine photodimers in cells from XPV patients.
Proc Natl Acad Sci U S A. 2009; 106(28):11552-7 [PubMed] Article available free on PMC after 12/11/2015 Related Publications
Human cells tolerate UV-induced cyclobutane pyrimidine dimers (CPD) by translesion DNA synthesis (TLS), carried out by DNA polymerase eta, the POLH gene product. A deficiency in DNA polymerase eta due to germ-line mutations in POLH causes the hereditary disease xeroderma pigmentosum variant (XPV), which is characterized by sunlight sensitivity and extreme predisposition to sunlight-induced skin cancer. XPV cells are UV hypermutable due to the activity of mutagenic TLS across CPD, which explains the cancer predisposition of the patients. However, the identity of the backup polymerase that carries out this mutagenic TLS was unclear. Here, we show that DNA polymerase zeta cooperates with DNA polymerases kappa and iota to carry out error-prone TLS across a TT CPD. Moreover, DNA polymerases zeta and kappa, but not iota, protect XPV cells against UV cytotoxicity, independently of nucleotide excision repair. This presents an extreme example of benefit-risk balance in the activity of TLS polymerases, which provide protection against UV cytotoxicity at the cost of increased mutagenic load.

Ben Rekaya M, Messaoud O, Talmoudi F, et al.
High frequency of the V548A fs X572 XPC mutation in Tunisia: implication for molecular diagnosis.
J Hum Genet. 2009; 54(7):426-9 [PubMed] Related Publications
Xeroderma pigmentosum (XP, OMIM 278700-278780) is a group of autosomal recessive diseases characterized by hypersensitivity to UV rays. There are seven complementation groups of XP (XPA to XPG) and XPV. Among them, the XP group C (XP-C) is the most prevalent type in Western Europe and in the United States. We report here on the clinical and genetic investigation of XP-C patients in 14 Tunisian families. As the XPC V548A fs X572 mutation has been identified in Algerian and Moroccan populations, Tunisian patients were first screened for this mutation by a direct sequencing of exon 9 of the XPC gene. All patients with a severe clinical form had this mutation, thus showing the homogeneity of the mutational spectrum of XPC in Tunisia. A potential founder effect was searched and confirmed by haplotype analysis. Taking into account the similarity of the genetic background, we propose a direct screening of this mutation as a rapid and cost-effective tool for the diagnosis of XP-C in North Africa.

Di Lucca J, Guedj M, Lacapère JJ, et al.
Variants of the xeroderma pigmentosum variant gene (POLH) are associated with melanoma risk.
Eur J Cancer. 2009; 45(18):3228-36 [PubMed] Related Publications
PURPOSE: Xeroderma pigmentosum variant (XPV) is a rare recessive autosomal genodermatosis predisposing to multiple early onset skin cancers, including melanoma. XPV results from mutations of the POLH gene that encodes a DNA translesion polymerase. In this work, we tested the hypothesis that POLH variants could be associated with melanoma risk.
EXPERIMENTAL DESIGN: A common non-synonymous POLH variant, c.1783A>G p.M595V, was genotyped in 1075 melanoma patients and in 1091 ethnic-matched controls from France. In addition, we searched for rare POLH variants by sequencing the entire coding sequence in 201 patients having a familial history of melanoma (n=123), sporadic multiple melanomas (n=65) and a melanoma associated with a skin carcinoma (n=13).
RESULTS: Overall, the c.1783G, p.595V allele was statistically associated with melanoma (respective allelic frequencies, 0.040 versus 0.022, P-value=1.17 x 10(-3), odds ratio (OR)=1.86 [1.27-2.71]), which was further confirmed by a meta-analysis including 274 patients and 174 matched controls from Italy (P-value=7.7 x 10(-4), OR=1.84 [1.29-2.63]). Interestingly, three non-synonymous POLH variants were identified in three patients (c.295G>A p.V99M, c.815T>C p.I272T and c.1745C>T p.S582L) which were absent in 352 chromosome controls from healthy subjects.
CONCLUSIONS: Besides severe deficiencies in translesion synthesis which are major risks factors for skin carcinomas and melanomas, less deleterious POLH variants could act as low penetrance melanoma predisposing alleles. The ongoing identification of genetic markers implied in skin cancer predisposition could help to identify high-risk subjects as targets for clinical follow-up. Replication studies in other populations are awaited to assess these data.

Faili A, Stary A, Delbos F, et al.
A backup role of DNA polymerase kappa in Ig gene hypermutation only takes place in the complete absence of DNA polymerase eta.
J Immunol. 2009; 182(10):6353-9 [PubMed] Related Publications
Patients with the variant form of xeroderma pigmentosum (XPV) syndrome have a genetic deficiency in DNA polymerase (Pol) eta, and display accordingly an increased skin sensitivity to UV light, as well as an altered mutation pattern of their Ig V genes in memory B cells, alteration that consists in a reduced mutagenesis at A/T bases. We previously suggested that another polymerase with a different mutation signature, Pol kappa, is used as backup for Ig gene hypermutation in both humans and mice in cases of complete Pol eta deficiency, a proposition supported in this study by the analysis of Pol eta x Pol kappa double-deficient mice. We also describe a new XPV case, in which a splice site mutation of the first noncoding exon results in a decreased mRNA expression, a mRNA that otherwise encodes a normal Pol eta protein. Whereas the Pol eta mRNA level observed in patient's fibroblasts is one-twentieth the value of healthy controls, it is only reduced to one-fourth of the normal level in activated B cells. Memory B cells from this patient showed a 50% reduction in A/T mutations, with a spectrum that still displays a strict Pol eta signature. Pol eta thus appears as a dominant enzyme in hypermutation, its presence precluding the use of a substitute enzyme even in conditions of reduced availability. Such a dominant behavior may explain the lack of Pol kappa signature in Ig gene mutations of some XPV patients previously described, for whom residual Pol eta activity might exist.

Bétous R, Rey L, Wang G, et al.
Role of TLS DNA polymerases eta and kappa in processing naturally occurring structured DNA in human cells.
Mol Carcinog. 2009; 48(4):369-78 [PubMed] Article available free on PMC after 12/11/2015 Related Publications
Accurate DNA replication during S-phase is fundamental to maintain genome integrity. During this critical process, replication forks frequently encounter obstacles that impede their progression. While the regulatory pathways which act in response to exogenous replication stress are beginning to emerge, the mechanisms by which fork integrity is maintained at naturally occurring endogenous replication-impeding sequences remains obscure. Notably, little is known about how cells replicate through special chromosomal regions containing structured non-B DNA, for example, G4 quartets, known to hamper fork progression or trigger chromosomal rearrangements. Here, we have investigated the role in this process of the human translesion synthesis (TLS) DNA polymerases of the Y-family (pol eta, pol iota, and pol kappa), specialized enzymes known to synthesize DNA through DNA damage. We show that depletion by RNA interference of expression of the genes for Pol eta or Pol kappa, but not Pol iota, sensitizes U2OS cells treated with the G4-tetraplex interactive compound telomestatin and triggers double-strand breaks in HeLa cells harboring multiple copies of a G-rich sequence from the promoter region of the human c-MYC gene, chromosomally integrated as a transgene. Moreover, we found that downregulation of Pol kappa only raises the level of DSB in HeLa cells containing either one of two breakage hotspot structured DNA sequences in the chromosome, the major break region (Mbr) of BCL-2 gene and the GA rich region from the far right-hand end of the genome of the Kaposi Sarcoma associated Herpesvirus. These data suggest that naturally occurring DNA structures are physiological substrates of both pol eta and pol kappa. We discuss these data in the light of their downregulation in human cancers.

Inui H, Oh KS, Nadem C, et al.
Xeroderma pigmentosum-variant patients from America, Europe, and Asia.
J Invest Dermatol. 2008; 128(8):2055-68 [PubMed] Article available free on PMC after 12/11/2015 Related Publications
Xeroderma pigmentosum-variant (XP-V) patients have sun sensitivity and increased skin cancer risk. Their cells have normal nucleotide excision repair, but have defects in the POLH gene encoding an error-prone polymerase, DNA polymerase eta (pol eta). To survey the molecular basis of XP-V worldwide, we measured pol eta protein in skin fibroblasts from putative XP-V patients (aged 8-66 years) from 10 families in North America, Turkey, Israel, Germany, and Korea. Pol eta was undetectable in cells from patients in eight families, whereas two showed faint bands. DNA sequencing identified 10 different POLH mutations. There were two splicing, one nonsense, five frameshift (3 deletion and 2 insertion), and two missense mutations. Nine of these mutations involved the catalytic domain. Although affected siblings had similar clinical features, the relation between the clinical features and the mutations was not clear. POLH mRNA levels were normal or reduced by 50% in three cell strains with undetectable levels of pol eta protein, indicating that nonsense-mediated message decay was limited. We found a wide spectrum of mutations in the POLH gene among XP-V patients in different countries, suggesting that many of these mutations arose independently.

Lee DH, Pfeifer GP
Translesion synthesis of 7,8-dihydro-8-oxo-2'-deoxyguanosine by DNA polymerase eta in vivo.
Mutat Res. 2008; 641(1-2):19-26 [PubMed] Article available free on PMC after 12/11/2015 Related Publications
7,8-Dihydro-8-oxo-2'-deoxyguanosine (8-oxo-dG) is one of the most common DNA lesions induced by oxidative stress. This lesion can be bypassed by DNA polymerase eta (Pol eta) using in vitro translesion synthesis (TLS) reactions. However, the role that Pol eta plays in vivo contributing to 8-oxo-dG mutagenesis remains unclear. To clarify the role of Pol eta in 8-oxo-dG mutagenesis, we have used an siRNA knockdown approach in combination with a supF shuttle vector (pSP189) which replicates in mammalian cells. The pSP189 plasmid was treated with methylene blue plus light (MBL), which produces predominantly 8-oxo-dG in DNA, and was then replicated in GM637 cells in presence of siRNA that knocks down the expression of Pol eta, or in XP-V cells, which lack functional Pol eta. The mutant frequencies were increased in the Pol eta siRNA knockdown cells and in XP-V cells relative to control, meaning that Pol eta plays an important role in preventing 8-oxo-dG mutagenesis. In the same system, knockdown of OGG1 also led to an increase in mutagenesis. Neither the type of mutations nor their distribution along the supF gene were significantly different between control and target specific siRNA-transfected cells (or XP-V cells) and were predominantly G to T transversions. These results show that Pol eta has an important role in error-free 8-oxo-dG lesion bypass and avoidance of oxidative stress-induced mutagenesis in vivo.

Dworaczek H, Xiao W
Xeroderma pigmentosum: a glimpse into nucleotide excision repair, genetic instability, and cancer.
Crit Rev Oncog. 2007; 13(2):159-77 [PubMed] Related Publications
Xeroderma pigmentosum (XP) is a rare DNA repair disorder characterized by extreme sensitivity to sunlight and severe predisposition to UV-induced skin cancer. Seven genes, ranging from XPA to XPG, are defective in XP. These genes are important components of the nucleotide excision repair (NER) system, which removes DNA damage induced by solar radiation, thereby preventing genome instability and carcinogenesis. In addition, XPV patients are defective in a translesion synthesis activity specialized in bypassing UV-induced lesions, and share symptoms with other XP patients. This review will focus on the evidence that elucidates the link between defective NER, genetic instability, and oncogenesis.

Sugasawa K
Xeroderma pigmentosum genes: functions inside and outside DNA repair.
Carcinogenesis. 2008; 29(3):455-65 [PubMed] Related Publications
Xeroderma pigmentosum (XP) is an autosomal recessive disease, which is characterized by susceptibility to ultraviolet light (UV)-induced skin cancer. Among eight genes so far identified as responsible for XP, XPA through XPG are involved in nucleotide excision repair of DNA damage induced by UV as well as various chemical carcinogens. Since this repair system removes a major UV photoproduct, the cyclobutane pyrimidine dimer, quite slowly from the global genome, this lesion must be accurately bypassed during replication by DNA polymerase eta, encoded by the XPV gene. Recent studies have revealed that each of these XP genes possesses additional functions, some of which are concerned with other DNA repair pathways and/or cellular DNA damage responses. Such differential functions not only explain clinical heterogeneity among different genetic complementation groups but also have implications for the promotion of carcinogenic processes in XP patients.

Smith JM, Stubbert LJ, Hamill JD, McKay BC
The contribution of transactivation subdomains 1 and 2 to p53-induced gene expression is heterogeneous but not subdomain-specific.
Neoplasia. 2007; 9(12):1057-65 [PubMed] Article available free on PMC after 12/11/2015 Related Publications
Two adjacent regions within the transactivation domain of p53 are sufficient to support sequence-specific transactivation when fused to a heterologous DNA binding domain. It has been hypothesized that these two subdomains of p53 may contribute to the expression of distinct p53-responsive genes. Here we have used oligonucleotide microarrays to identify transcripts induced by variants of p53 with point mutations within subdomains 1, 2, or 1 and 2 (QS1, QS2, and QS1/QS2, respectively). The expression of 254 transcripts was increased in response to wild-type p53 expression but most of these transcripts were poorly induced by these variants of p53. Strikingly, a number of known p53-regulated transcripts including TNFRSF10B, BAX, BTG2, and POLH were increased to wild-type levels by p53(QS1) and p53(QS2) but not p53(QS1/QS2), indicating that either subdomain 1 or 2 is sufficient for p53-dependent expression of a small subset of p53-responsive genes. Unexpectedly, there was no evidence for p53(QS1)- or p53(QS2)-specific gene expression. Taken together, we found heterogeneity in the requirement for transactivation subdomains 1 and 2 of p53 without any subdomain-specific contribution to p53-induced gene expression.

Armelini MG, Lima-Bessa KM, Marchetto MC, et al.
Exploring DNA damage responses in human cells with recombinant adenoviral vectors.
Hum Exp Toxicol. 2007; 26(11):899-906 [PubMed] Related Publications
Recombinant adenoviral vectors provide efficient means for gene transduction in mammalian cells in vitro and in vivo. We are currently using these vectors to transduce DNA repair genes into repair deficient cells, derived from xeroderma pigmentosum (XP) patients. XP is an autosomal syndrome characterized by a high frequency of skin tumors, especially in areas exposed to sunlight, and, occasionally, developmental and neurological abnormalities. XP cells are deficient in nucleotide excision repair (affecting one of the seven known XP genes, xpa to xpg) or in DNA replication of DNA lesions (affecting DNA polymerase eta, xpv). The adenovirus approach allows the investigation of different consequences of DNA lesions in cell genomes. Adenoviral vectors carrying several xp and photolyases genes have been constructed and successfully tested in cell culture systems and in vivo directly in the skin of knockout model mice. This review summarizes these recent data and proposes the use of recombinant adenoviruses as tools to investigate the mechanisms that provide protection against DNA damage in human cells, as well as to better understand the higher predisposition of XP patients to cancer.

Tanioka M, Masaki T, Ono R, et al.
Molecular analysis of DNA polymerase eta gene in Japanese patients diagnosed as xeroderma pigmentosum variant type.
J Invest Dermatol. 2007; 127(7):1745-51 [PubMed] Related Publications
POLH mutations were identified in 16 Japanese patients, who were diagnosed, both clinically and at a cellular level, as being of the xeroderma pigmentosum variant type (XPV). While all the patients developed skin cancer with an average onset of the cancer at 45 years, in non-XP Japanese the onset was at over 70 years. All the cell strains from the patients were normal or slightly hypersensitive to UV and most of these showed enhanced UV sensitivity when the post-UV colony formation was performed in the presence of caffeine. Immunoprecipitation analysis with two kinds of anti-POLH protein antibodies revealed that cells from 13 patients did not show the 83 kDa POLH band and that cells from one patient had a faint 83 kDa band. All of these 14 cell strains, without a POLH band or with a weak POLH band, had mutations in the POLH gene. The IP analysis of the POLH protein revealed a very useful method for screening the patients suspected of XPV. Seven mutations in the POLH gene including three novel mutations were identified. Among the mutations detected, 11 alleles out of 28 (39%) were G490T mutations.

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