Hydrogen peroxide induced genomic instability in nucleotide excision repair-deficient lymphoblastoid cells

Copyright @ 2010 Gopalakrishnan et al; licensee BioMed Central Ltd.Background The Nucleotide Excision Repair (NER) pathway specialises in UV-induced DNA damage repair. Inherited defects in the NER can predispose individuals to Xeroderma Pigmentosum (XP). UV-induced DNA damage cannot account for the manifestation of XP in organ systems not directly exposed to sunlight. While the NER has recently been implicated in the repair of oxidative DNA lesions, it is not well characterised. Therefore we sought to investigate the role of NER factors Xeroderma Pigmentosum A (XPA), XPB and XPD in oxidative DNA damage-repair by subjecting lymphoblastoid cells from patients suffering from XP-A, XP-D and XP-B with Cockayne Syndrome to hydrogen peroxide (H2O2). Results Loss of functional XPB or XPD but not XPA led to enhanced sensitivity towards H2O2-induced cell death. XP-deficient lymphoblastoid cells exhibited increased susceptibility to H2O2-induced DNA damage with XPD showing the highest susceptibility and lowest repair capacity. Furthermore, XPB- and XPD-deficient lymphoblastoid cells displayed enhanced DNA damage at the telomeres. XPA- and XPB-deficient lymphoblastoid cells also showed differential regulation of XPD following H2O2 treatment. Conclusions Taken together, our data implicate a role for the NER in H2O2-induced oxidative stress management and further corroborates that oxidative stress is a significant contributing factor in XP symptoms. Resistance of XPA-deficient lymphoblastoid cells to H2O2-induced cell death while harbouring DNA damage poses a potential cancer risk factor for XPA patients. Our data implicate XPB and XPD in the protection against oxidative stress-induced DNA damage and telomere shortening, and thus premature senescence.This research is supported by the Defence Innovative Research Programme, Defence Science and Technology Agency, Singapore (POD: 0613592) and the Academic Research Fund, Ministry of Education, Singapore (T206B3108). Supported in part by a grant from British Council, PMI2 Connect (Grant Number: RC134)

GENE EXPRESSION KINETICS AND PROTEIN DISTRIBUTION OF NUCLEOTIDE EXCISION REPAIR FACTORS IN THE INNER EAR AS A FUNCTION OF cis-DIAMMINEDICHLOROPLATINUM-II DNA DAMAGE

The kinetics of the rate-limiting genes of the molecular DNA repair pathways of nucleotide excision repair (NER) were quantified from the inner ear as a function of cis-diamminedichloroplatinum-II (cisplatin) treatment. The distribution of the post-translational products of these genes was evaluated among neurons and sensory hair cells of the inner ear following cisplatin treatment. These NER factors (genes & post-translational products) are only potentiated by DNA damage and are particularly sensitive to cisplatin induced DNA damage. A 2 x 3 x 2 factorial design, consisting of two treatment conditions (saline and cisplatin treated Fischer344 rats), three survival times and two molecular analysis methods (polymerase chain reaction and immunohistochemistry) was employed in this dissertation. The results revealed at least five important findings. First, it revealed for the first time that complex DNA repair molecular pathways such as NER exist in the inner ear. Second, it revealed for the first time that molecules used by advanced tumor cells to detect and repair damaged DNA from cisplatin genotoxicity also generalize to the inner ear and are stimulated by even small sub-toxic doses of cisplatin. Third, it revealed for the first time that NER proteins reside in the cytoplasm of neurons under normal conditions and translocate to the nucleus under conditions of genomic stress. Fourth, it revealed for the first time that the basal coil of the mammalian cochlea differs from the apical coil in the magnitude and latency in which NER molecules translocate from the cytoplasm to the nucleus under conditions of genomic stress. Fifth, the current work provides the bases for a new line of hearing research focused on molecular mechanisms of inner ear DNA repair

ING1 and ING2: Multi-Faceted Tumor Suppressor Genes

International audienceING1 (Inhibitor of Growth 1) was identified and characterized as a "candidate" tumor suppressor gene in 1996. Subsequently four more genes, also characterized as "candidate" tumor suppressor genes, were identified by homology search: ING2, ING3, ING4 and ING5. The ING proteins are characterized by a high homology in their C-terminal domain which contains a Nuclear Localization Sequence (NLS) and a Plant HomeoDomain (PHD) which has a high affinity to Histone 3 tri-methylated on lysine 4 (H3K4Me3). The ING proteins have been involved in the control of cell growth, senescence, apoptosis, chromatin remodelling and DNA repair. Within the ING family, ING1 and ING2 form a subgroup since they are evolutionarily and functionally close. In Yeast, only one gene, Pho23, is related to ING1 and ING2 and possesses also a PHD. Recently, the ING1 and ING2 tumor suppressor status has been fully established since several studies have described the loss of ING1 and ING2 protein expression in human tumors and both ING1 and ING2 knockout mice were reported to have spontaneously developed tumors, B-cell lymphomas and soft tissue sarcomas respectively. In this review we will describe for the first time what is known about ING1 and ING2 genes, proteins, their regulations in both human and mice, and their status in human tumors. Furthermore, we explore the current knowledge about identified functions involving ING1 and ING2 in tumor suppression pathways especially in the control of cell cycle and in genome stability

Untersuchungen zum Einfluss von Sulforaphan auf die DNA-Einzelstrangbruch- und Nukleotidexzisionsreparatur

Eine Risikobewertung von Broccoli-basierten Nahrungsergänzungsmitteln fehlt bis dato. Die vorliegende Arbeit leistet in diesem Kontext nun erstmals einen kritischen Beitrag zur Auswirkung der bioaktiven Substanz Sulforaphan (SFN) auf essentielle DNA-Reparaturprozesse, die anhand der Prozessierung und Reparatur zweier Modellläsionen in Zellkulturen beurteilt wurden. Es zeigte sich, dass SFN beide Reparaturwege inhibiert und Zink-bindende Strukturen dabei potentielle Angriffspunkte darstellen

Developing small molecule inhibitors targeting Replication Protein A for platinum-based combination therapy

Indiana University-Purdue University Indianapolis (IUPUI)All platinum (Pt)-based chemotherapeutics exert their efficacy primarily via the formation of DNA adducts which interfere with DNA replication, transcription and cell division and ultimately induce cell death. Repair and tolerance of Pt-DNA lesions by nucleotide excision repair and homologous recombination (HR) can substantially reduce the effectiveness of the Pt therapy. Inhibition of these repair pathways, therefore, holds the potential to sensitize cancer cells to Pt treatment and increase clinical efficacy. Replication Protein A (RPA) plays essential roles in both NER and HR, along with its role in DNA replication and DNA damage checkpoint activation. Each of these functions requires RPA binding to single-stranded DNA (ssDNA). We synthesized structural analogs of our previously reported RPA inhibitor TDRL-505, determined the structure activity relationships and evaluated their efficacy in tissue culture models of epithelial ovarian cancer (EOC) and non-small cell lung cancer (NSCLC). These data led us to the identification of TDRL-551, which exhibited a greater than 2-fold increase in in vitro and cellular activity. TDRL-551 showed synergy with Pt in tissue culture models of EOC and in vivo efficacy, as a single agent and in combination with platinum, in a NSCLC xenograft model. These data demonstrate the utility of RPA inhibition in EOC and NSCLC and the potential in developing novel anticancer therapeutics that target RPA-DNA interactions

Role of Xeroderma pigmentosum complementation group B (XPB) in genome maintenance under oxidative stress

Master'sMASTER OF SCIENC