Different Effects ofCSAandCSBDeficiency on Sensitivity to Oxidative DNA Damage

Abstract

Mutations in the CSA and CSB genes cause Cockayne syndrome, a rare inherited disorder characterized by UV sensitivity, severe neurological abnormalities, and progeriod symptoms. Both gene products function in the transcription-coupled repair (TCR) subpathway of nucleotide excision repair (NER), providing the cell with a mechanism to remove transcription-blocking lesions from the transcribed strands of actively transcribed genes. Besides a function in TCR of NER lesions, a role of CSB in (transcription-coupled) repair of oxidative DNA damage has been suggested. In this study we used mouse models to compare the effect of a CSA or a CSB defect on oxidative DNA damage sensitivity at the levels of the cell and the intact organism. In contrast to CSB ؊/؊ mouse embryonic fibroblasts (MEFs), CSA ؊/؊ MEFs are not hypersensitive to gamma-ray or paraquat treatment. Similar results were obtained for keratinocytes. In contrast, both CSB ؊/؊ and CSA ؊/؊ embryonic stem cells show slight gamma-ray sensitivity. Finally, CSB ؊/؊ but not CSA ؊/؊ mice fed with food containing di(2-ethylhexyl)phthalate (causing elevated levels of oxidative DNA damage in the liver) show weight reduction. These findings not only uncover a clear difference in oxidative DNA damage sensitivity between CSA-and CSB-deficient cell lines and mice but also show that sensitivity to oxidative DNA damage is not a uniform characteristic of Cockayne syndrome. This difference in the DNA damage response between CSA-and CSBdeficient cells is unexpected, since until now no consistent differences between CSA and CSB patients have been reported. We suggest that the CSA and CSB proteins in part perform separate roles in different DNA damage response pathways.

The role of CSA in the response to oxidative DNA damage in human cells

“…Whether CSA is involved in oxidative DNA damage response is still an open question. In contrast to CS-B cells, CS-A mouse fibroblasts did not show hypersensitivity to either IR or paraquat (de Waard et al, 2004), suggesting that CSA might not be involved in oxidative DNA damage sensitivity. On the other hand, a recent study indicates that the processing of plasmids containing a single 8-OH-Gua results in similarly defective host-cell reactivation in CS-A as well as in CS-B cell lines (Spivak and Hanawalt, 2006).…”

The role of Cockayne Syndrome group B (CSB) protein in base excision repair and aging

“…Importantly, after exposure to IR, CSB deficient transformed fibroblasts, mouse embryonic fibroblasts (MEF), embryonic stem (ES) cells and keratinocytes from CSB knockout mice all show a marked reduction in survival (de Waard et al, 2004;de Waard et al, 2003;Leadon and Cooper, 1993;Tuo et al, 2001). While IR induces a variety of DNA lesions including single stranded DNA breaks (SSBs), double-strand DNA breaks (DSBs) and oxidative base damage, the observed hypersensitivity has been ascribed to oxidative DNA modifications (de Waard et al, 2004;de Waard et al, 2003), which normally are repaired by BER.…”

“…While IR induces a variety of DNA lesions including single stranded DNA breaks (SSBs), double-strand DNA breaks (DSBs) and oxidative base damage, the observed hypersensitivity has been ascribed to oxidative DNA modifications (de Waard et al, 2004;de Waard et al, 2003), which normally are repaired by BER. Indeed, after IR treatment, CSB deficient primary and transformed fibroblasts, and cells with mutated ATPase domains V (CSBT912V and CSBT912/913V) and VI (CSBQ942E and CSBR946A) accumulated significant amounts of the oxidative base modifications 8-hydroxyguanine (8-oxoG) and 8-hydroxyadenine (8-oxoA) (Tuo et al, 2003;Tuo et al, 2002b).…”

“…Enzymatic reduction of paraquat produces paraquat radicals (PQ + ), which react with molecular oxygen to generate reactive oxygen species (ROS) in several tissues inflicting oxidative stress to cells and thereby causes oxidative DNA lesions (Ali et al, 1996). CSB knockout MEFs were found also to be hypersensitive to paraquat (de Waard et al, 2004;de Waard et al, 2003). Potassium bromate (KBrO 3 ) generally acts mostly as a oneelectron oxidant that leads almost exclusively to the formation of 8-oxoG and formamidopyrimidine derivatives.…”

Clinical implications of the basic defects in Cockayne syndrome and xeroderma pigmentosum and the DNA lesions responsible for cancer, neurodegeneration and aging

“…The neurological symptoms of CS patients have been ascribed to defective repair in the brain of endogenous oxidative damage that blocks transcription (Kuraoka et al, 2000;Osterod et al, 2002;de Waard et al, 2003;Kyng et al, 2003;Tuo et al, 2003;Cline et al, 2004), but the more common oxidative base damages (8-oxo-G, 5-hydroxycytosine, thymine glycols) do not block transcription and are therefore not the culprits in neurodegeneration (Kathe et al, 2004). CSA and CSB cells are however different in their responses to oxidative damage, despite overlap in clinical symptoms (de Waard et al, 2004;D'Errico et al, 2007). The oxidative lesion 8-oxo-G does not appear to accumulate in CS autopsy material (Hayashi et al, 2005) despite the higher amounts of protein oxidation and lipid oxidation in the brains of CS patients (Hayashi et al, 2001) and crossing Cs-b mice with Ogg1 deficient mice did not enhance the neurological symptoms (Laposa et al, 2007b).…”