Nucleotide excision repair is a highly versatile DNA repair system responsible for elimination of a wide variety of lesions from the genome. It is comprised of two subpathways: transcription-coupled repair that accomplishes efficient removal of damage blocking transcription and global genome repair. Recently, the basic mechanism of global genome repair has emerged from biochemical studies. However, little is known about transcription-coupled repair in eukaryotes. Here we report the identification of a novel protein designated XAB2 (XPA-binding protein 2) that was identified by virtue of its ability to interact with XPA, a factor central to both nucleotide excision repair subpathways. The XAB2 protein of 855 amino acids consists mainly of 15 tetratricopeptide repeats. In addition to interacting with XPA, immunoprecipitation experiments demonstrated that a fraction of XAB2 is able to interact with the transcription-coupled repair-specific proteins CSA and CSB as well as RNA polymerase II. Furthermore, antibodies against XAB2 inhibited both transcriptioncoupled repair and transcription in vivo but not global genome repair when microinjected into living fibroblasts. These results indicate that XAB2 is a novel component involved in transcription-coupled repair and transcription.

Transcription-coupled repair (TCR) efficiently removes a variety of lesions from the transcribed strand of active genes. By allowing rapid resumption of RNA synthesis, the process is of major importance for cellular resistance to transcription-blocking genotoxic damage. Mutations in the Cockayne syndrome group A or B (CSA or CSB) gene result in defective TCR. However, the exact mechanism of TCR in mammalian cells remains to be elucidated. We found that CSA protein is rapidly translocated to the nuclear matrix after UV irradiation. The translocation of CSA was independent of Xeroderma pigmentosum group C, which is specific to the global genome repair subpathway of nucleotide excision repair (NER) and of the core NER factor Xeroderma pigmentosum group A but required the CSB protein. In UV-irradiated cells, CSA protein colocalized with the hyperphosphorylated form of RNA polymerase II, engaged in transcription elongation. The translocation of CSA was also induced by treatment of the cells with cisplatin or hydrogen peroxide, both of which produce damage that is subjected to TCR but not induced by treatment with dimethyl sulfate, which produces damage that is not subjected to TCR. The hydrogen peroxide-induced translocation of CSA was also CSB dependent. These findings establish a link between TCR and the nuclear matrix mediated by CSA.

Diabetes mellitus is known to exacerbate cerebral ischemic injury. In the present study, we investigated antiapoptotic and anti-inflammatory effects of oral supplementation of ascorbic acid (AA) on cerebral injury caused by middle cerebral artery occlusion and reperfusion (MCAO/Re) in rats with streptozotocin-induced diabetes. We also evaluated the effects of AA on expression of sodium-dependent vitamin C transporter 2 (SVCT2) and glucose transporter 1 (GLUT1) after MCAO/Re in the brain. The diabetic state markedly aggravated MCAO/Re-induced cerebral damage, as assessed by infarct volume and edema. Pretreatment with AA (100 mg/kg, p.o.) for two weeks significantly suppressed the exacerbation of damage in the brain of diabetic rats. AA also suppressed the production of superoxide radical, activation of caspase-3, and expression of proinflammatory cytokines (tumor necrosis factor-α and interleukin-1β) in the ischemic penumbra. Immunohistochemical staining revealed that expression of SVCT2 was upregulated primarily in neurons and capillary endothelial cells after MCAO/Re in the nondiabetic cortex, accompanied by an increase in total AA (AA + dehydroascorbic acid) in the tissue, and that these responses were suppressed in the diabetic rats. AA supplementation to the diabetic rats restored these responses to the levels of the nondiabetic rats. Furthermore, AA markedly upregulated the basal expression of GLUT1 in endothelial cells of nondiabetic and diabetic cortex, which did not affect total AA levels in the cortex. These results suggest that daily intake of AA attenuates the exacerbation of cerebral ischemic injury in a diabetic state, which may be attributed to anti-apoptotic and anti-inflammatory effects via the improvement of augmented oxidative stress in the brain. AA supplementation may protect endothelial function against the exacerbated ischemic oxidative injury in the diabetic state and improve AA transport through SVCT2 in the cortex.

BackgroundGanoderma lucidum is a popular medicinal mushroom used for promoting health and longevity in Asian countries. Previously, we reported that a water-soluble extract from a culture medium of Ganoderma lucidum mycelia (MAK) exerts antioxidative and cerebroprotective effects against ischemia–reperfusion injury in vivo. Here, we evaluated the antidepressant and anxiolytic activities of MAK in rats.MethodsMAK (0.3 or 1 g/kg, p.o.) was administered in the experimental animals 60 min before the forced swimming, open-field, elevated plus-maze, contextual fear-conditioning, and head twitch tests. Additionally, the mechanisms involved in the antidepressant-like action of MAK were investigated by the serotonin precursor 5-hydroxy-L-tryptophan (5-HTP)- or 5-HT2A agonist (±)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane hydrochloride (DOI)-induced head twitch responses.ResultsTreatment with MAK (1 g/kg) exhibited antidepressant-like effects in the forced swimming test, attenuated freezing behavior in the contextual fear-conditioning test, and decreased the number of head twitches induced by DOI, but not with 5-HTP. No significant response was observed in locomotion or anxiety-like behavior, when the animals were evaluated in the open-field or elevated plus-maze test, respectively.ConclusionsThese data suggest that MAK has antidepressant-like potential, which is most likely due to the antagonism of 5-HT2A receptors, and possesses anxiolytic-like effects toward memory-dependent and/or stress-induced anxiety in rats.