DNA excision-repair defect of xeroderma pigmentosum prevents removal of a class of oxygen free radical-induced base lesions.

Abstract

Plasmid DNA was rirradiated or treated with H202 in the presence of Cu2+ to generate oxygen free radical-induced lesions. Open circular DNA molecules were removed by ethidium bromide/CsCI density gradient centrifugation. The closed circular DNA fraction was treated with the Escherichia coli reagent enzymes endonuclease IH (Nth protein) and Fpg protein. This treatment converted DNA molecules containing the major base lesions pyrimidine hydrates and 8-hydroxyguanine to a nicked form. Remaining closed circular DNA containing other oxygen radical-induced base lesions was used as a substrate for nudeotide excision-repair in a cell-free system. Extracts from normal human cells, but not extracts from xeroderma pigmentosum cells, catalyzed repair synthesis in this DNA. The repair defect in the latter extracts could be specifically corrected by in vitro complementation. The data suggest that accumulation of endogenous oxidative damage in cellular DNA from xeroderma pigmentosum patients contributes to the increased frequency ofinternal cancers and the neural degeneration occurring in serious cases of the syndrome.UV light and ionizing radiation induce the formation of different and diverse types of DNA lesions. After exposure of DNA to UV light, pyrimidine dimers are the main products, whereas oxygen free radicals produced by ionizing radiation generate single-and double-strand breaks as well as a variety of ring-saturated, ring-fragmented, and oxidized base derivatives. Different DNA repair processes are used by cells to correct the damage: pyrimidine dimers are removed as part of oligonucleotides in a nucleotide excision-repair pathway initiated by a multisubunit incision nuclease, while several oxidized bases are liberated in free form by DNA glycosylases in a base excision-repair process. In cells from patients with the human inherited disease xeroderma pigmentosum (XP), incision at pyrimidine dimers is defective. As a consequence, the main symptoms of XP involve skin and eye lesions occurring after exposure to sunlight, including multiple skin cancers. However, other symptoms of XP are not readily explained as being due to UV-induced DNA damage. In severe cases, sometimes termed de SanctisCacchione syndrome, patients exhibit progressive neurological deterioration with massive loss of neurons and a 10-to 20-fold increase in frequency of several types of internal cancers (1). The origins of these symptoms have been unclear, but they could arise because of accumulation of DNA damage generated by exposure to environmental chemical mutagens or reactive cellular metabolites (2). In the latter regard, DNA is known to undergo endogenous damage due to hydrolysis, reaction with active oxygen, and nonenzymatic methylation. However, the main known lesions introduced in this fashion are corrected by base excision-repair (3).Reagent Enzymes and DNA Substrates. The following proteins were purified from Escherichia coli overproducer strains as described: E. coli endonuclease III (Nth protein) (4), E. coli Fpg protein (5), a...

Identification and Quantification of a Guanine−Thymine Intrastrand Cross-Link Lesion Induced by Cu(II)/H2O2/Ascorbate

“…Along this line, it was shown that repair-defective cells from severe cases of XP appear to be incapable of removing a major type of oxidative DNA damage (61). ROS may generate abundant DNA intrastrand crosslink lesions in the central nervous system because neurons consume a large amount of oxygen.…”

1-Methyl-4-phenyl-pyridinium time-dependently alters expressions of oxoguanine glycosylase 1 and xeroderma pigmentosum group F protein in PC12 cells

“…It is commonly known that DNA oxidative damage is repaired via activation of both BER and NER pathways, based on the severity of DNA damage [13][14][15][16]29] . Theoretically, DNA repair function should be activated once DNA damage occurs in the cells.…”

“…It has been shown that OGG1 activity is inversely related to the level of 8-oxodG in brain regions [12] . Alternatively, oxidative DNA damage can be repaired by nucleotide excision repair (NER) [13][14][15][16][17][18] , which is a major DNA repair pathway in removing a wide range of DNA damage including DNA double helix distortion [19] . Approximately 20-30 proteins are involved in NER, participating in successive steps of damage recognition, local opening of the DNA double helix around the injury, incision of the damaged strand on either side of the lesion, and nascent DNA synthesis/ligation [19] .…”

Developing master keys to brain pathology, cancer and aging from the structural biology of proteins controlling reactive oxygen species and DNA repair

“…These typically occur from carcinogenic compounds and covalent linkages between pyrimidine bases caused by UV exposure. Clearly, UV light does not penetrate the skull, but there is evidence that DNA exposed to oxygen radicals generated by ionizing radiation can produce DNA lesions that require NER for repair (Satoh et al, 1993). NER is a particularly versatile DNA repair system that is capable of global genome repair (GG-NER) and also transcription-coupled repair (TC-NER), where the template strand of actively transcribed genes is preferribly repaired.…”