XRCC1 participates in DNA single strand break and base excision repair (BER) to preserve genetic stability in mammalian cells. XRCC1 participation in these pathways is mediated by its interactions with several of the acting enzymes. Here, we report that XRCC1 interacts physically and functionally with hOGG1, the human DNA glycosylase that initiates the repair by BER of the mutagenic oxidized base 8-oxoguanine. This interaction leads to a 2-to 3-fold stimulation of the DNA glycosylase activity of hOGG1. XRCC1 stimulates the formation of the hOGG1 Schiff-base DNA intermediate without interfering with the endonuclease activity of APE1, the second enzyme in the pathway. On the contrary, the stimulation in the appearance of the incision product seems to reflect the addition of the effects of XRCC1 on the two first enzymes of the pathway. The data presented support a model by which XRCC1 will pass on the DNA intermediate from hOGG1 to the endonuclease APE1. This results in an acceleration of the overall repair process of oxidized purines to yield an APE1-cleaved abasic site, which can be used as a substrate by DNA polymerase . More importantly, the results unveil a highly coordinated mechanism by which XRCC1, through its multiple protein-protein interactions, extends its orchestrating role from the base excision step to the resealing of the repaired DNA strand.A major threat to genetic stability is the damaging of DNA by either endogenous or exogenous compounds. This is underscored by the cancer-prone phenotype of human cells defective in DNA repair processes. Exposure of the cellular DNA to reactive oxygen species (ROS), 1 generated either by the normal metabolism of the cell or by chemical and physical exogenous agents, is at the origin of lesions that can have genotoxic or mutagenic consequences. To avoid the effects of ROS and, therefore, to maintain the integrity of their genetic information, organisms have evolved multiple DNA repair mechanisms (1). Because of its capacity to pair with an adenine during replication, 7,8-dihydro-8-oxoguanine (8-oxoG), an oxidized derivative of guanine, is arguably the major mutagenic lesion in DNA. Indeed, in Escherichia coli, the inactivation of the genes involved in the repair of this oxidized base leads to one of the strongest spontaneous mutator phenotypes, characterized by the exclusive increase in G to T transversions. Like for other ROS-induced modifications of DNA, 8-oxoG is mainly repaired by the base excision repair (BER) pathway. This pathway is initiated by the recognition and excision of the oxidized guanine by a DNA glycosylase, OGG1 being the major one in yeast and mammalian cells (2). In human cells, the resulting abasic (apurinic/apyrimidinic (AP)) site can be cleaved by a second enzymatic activity of the hOGG1 polypeptide, namely an AP lyase activity. If such a reaction takes place, the nick produced has a 3Ј-open aldehyde residue that is supposed to be removed by the 3Ј-deoxyribose phosphatase activity of APE1, the major AP endonuclease. However, recent data s...
In adults, stem cells are responsible for the maintenance of many actively renewing tissues, such as haematopoietic, skin, gut and germinal tissues. These stem cells can self-renew or be committed to becoming progenitors. Stem-cell commitment is thought to be irreversible but in male and female Drosophila melanogaster, it was shown recently that differentiating germ cells can revert to functional stem cells that can restore germinal lineage. Whether progenitors are also able to generate stem cells in mammals remains unknown. Here we show that purified mouse spermatogonial progenitors committed to differentiation can generate functional germinal stem cells that can repopulate germ-cell-depleted testes when transplanted into adult mice. We found that GDNF, a key regulator of the stem-cell niche, and FGF2 are able to reprogram in vitro spermatogonial progenitors for reverse differentiation. This study supports the emerging concept that the stem-cell identity is not restricted in adults to a definite pool of cells that self-renew, but that stemness could be acquired by differentiating progenitors after tissue injury and throughout life.
The Cockayne syndrome B (CSB) gene product is involved in the repair of various types of base modifications in actively transcribed DNA sequences. To investigate its significance for the repair of endogenous oxidative DNA damage, homozygous csb 7/7 /ogg1 7/7 double knockout mice were generated. These combine the deficiency of CSB with that of OGG1, a gene coding for the mammalian repair glycosylase that initiates the base excision repair of 7,8-dihydro-8-oxoguanine (8-oxoG). Compared to ogg1 7/7 mice, csb 7/7 /ogg1 7/7 mice were found to accumulate with age severalfold higher levels of oxidited purine modifications in hepatocytes, splenocytes and kidney cells. In contrast, the basal (steady-state) levels of oxidative DNA modifications in cells from csb 7/7 mice were not different from those in wild-type mice and did not increase with age. The analysis of the repair rates of additional oxidative DNA base modifications induced by photosensitization in immortalized embryonic fibroblasts was in accordance with these findings: compared to wild-type cells, the global repair was only slightly affected in csb 7/7 cells, more compromised in ogg1 7/7 cells, but virtually absent in csb 7/7 /ogg1 7/7 cells. An inhibition of transcription by a-amanitin did not block the Csbdependent repair in ogg1 7/7 fibroblasts. The influence of Csb on the global repair of 8-oxoG was not detectable in assays with total protein extracts and in a shuttle vector system. The data indicate a role for Csb in the removal of 8-oxoG from the overall genome that is independent of both Ogg1-mediated base excision repair and regular transcription.
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