Papel da proteína de reparo XPC na regulação das proteínas de reparo APE1, OGG1 e PARP-1 em células humanas e de camundongos

studies using UV as a source of DNA damage. However, even though unrepaired UV-induced DNA damages are related to mutagenesis, cell death and tumorigenesis, they do not explain phenotypes such as neurodegeneration and internal tumors observed in patients with syndromes like Xeroderma Pigmentosum (XP) and Cockayne Syndrome (CS) that are associated with NER deficiency. Recent evidences point to a role of NER in the repair of 8-oxodG, a typical substrate of Base Excision Repair (BER). Since deficiencies in BER result in genomic instability, neurodegenerative diseases and cancer, it was investigated in this research the impact of XPC deficiency on BER functions in human cells. It was analyzed both the expression and the cellular localization of APE1, OGG1 e PARP-1, the mainly BER enzymes, in different NER-deficient human fibroblasts. The endogenous levels of these enzymes are reduced in XPC deficient cells. Surprisingly, XP-C fibroblasts were more resistant to oxidative agents than the other NER deficient fibroblasts, despite presenting the highest of 8-oxodG. Furthermore, subtle changes in the nuclear and mitochondrial localization of APE1 were detected in XP-C fibroblasts. To confirm the impact of XPC deficiency in the regulation of APE1 and OGG1 expression and activity, we constructed a XPC-complemented cell line. Although the XPC complementation was only partial, we found that XPC-complemented cells presented increased levels of OGG1 than XPC-deficient cells. The extracts from XPC-complemented cells also presented an elevated OGG1 enzimatic activity. However, it was not observed changes in APE1 expression and activity in the XPCcomplemented cells. In addition, we found that full-length APE1 (37 kDa) and OGG1- α are in the mitochondria of XPC-deficient fibroblasts and XPC-complemented fibroblasts before and after induction of oxidative stress. On the other hand, the expression of APE1 and PARP-1 are not altered in brain and liver of XPC knockout mice. However, XPC deficiency changed the APE1 localization in hypoccampus and hypothalamus. We also observed a physical interaction between XPC and APE1 proteins in human cells. In conclusion, the data suggest that XPC protein has a role in the regulation of OGG1 expression and activity in human cells and is involved mainly in the regulation of APE1 localization in mice. Aditionally, the response of NER deficient cells under oxidative stress may not be only associated to the NER deficiency per se, but it may include the new functions of NER enzymes in regulation of expression and cell localization of BER proteins

Associação entre polimorfismos em enzimas de reparo de DNA e ocorrência de meningite bacteriana.

Despite advances in vaccine development and therapy, bacterial meningitis (BM) remains a major cause of death and long-term neurological disabilities. As part of the host inflammatory response to the invading pathogen, factors such as reactive oxygen species are generated, which may damage DNA and trigger the overactivation of DNA repair mechanisms. It is conceivable that the individual susceptibility and outcome of BM may be in part determined by non synonymous polymorphisms that may alter the function of crucial BER DNA repair enzymes as PARP-1, OGG-1 and APE-1. These enzymes, in addition to their important DNA repair function, also perform role of inflammatory regulators. In this work was investigated the non synonymous SNPs APE-1 Asn148Glu, OGG-1 Ser326Cys,PARP-1 Val762Ala, PARP-1 Pro882Leu and PARP-1 Cys908Tyr in patients with bacterial meningitis (BM), chronic meningitis (CM), aseptic meningitis (AM) and not infected (controls). As results we found increased frequency of variant alleles of PARP-1 Val762Ala (P = 0.005) and APE-1 Asn148Glu (P=0.018) in BM patients, APE-1 Asn148Glu in AM patients (P = 0.012) and decrease in the frequency of the variant allele OGG-1 Ser326Cys in patients with CM (P = 0.013), regarding the allelic frequencies in the controls. A major incidence of individuals heterozygous and/ or polymorphic homozygous in BM for PARP-1 Val762Ala (P= 0.0399, OD 4.2, 95% IC 1.213 -14.545) and PARP-1 Val762Ala/ APE-1 Asn148Glu (P = 0.0238, OD 11.111, 95% IC 1.274 - 96.914) was observed related to what was expected in a not infected population. It was also observed a major incidence of combined SNPs in the BM patients compared with the control group (P=0.0281), giving evidences that SNPs can cause some susceptibility to the disease. This combined effect of SNPs seems to regulate the principal cytokines and other factors related to BM inflammatory response and point the importance of DNA repair not only to repair activity when DNA is damaged, but to others essential functions to human organism balance.

Association of the XPD and XRCC3 gene polymorphisms with oral squamous cell carcinoma in a Northeastern Brazilian population: A pilot study

Objective to evaluate the association between XPD and XRCC3 polymorphisms and oral squamous cell carcinoma (OSCC). Design the sample consisted of 54 cases of OSCC and 40 cases of inflammatory fibrous hyperplasia (IFH). Genotypes were determined by the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. Results XPD-Lys/Gln was more common in IFH (n = 28; 70%) than in OSCC (n = 24; 44.4%) (OR: 0.3; p < 0.05). XPD-Gln was more frequent in high-grade lesions (0.48) than in low-grade lesions (0.21) (OR: 3.4; p < 0.05). The Gln/Gln genotype was associated with III and IV clinical stages (OR: 0.07; p < 0.05). XRCC3-Met was more frequent in OSCC (0.49) than in IFH (0.35) (OR: 2.6; p < 0.05). The Met/Met genotype was associated with the presence of metastases (OR: 8.1; p < 0.05) and with III and IV clinical stages (OR: 0.07; p < 0.05). Conclusions in this sample, the frequency of XPD-Gln in IFH suggests that this variant may protect against OSCC. The presence of the XRCC3-Met allele seems to contribute to the development of OSCC, metastases and more advanced stages in these lesions.

Association of the XPD and XRCC3 gene polymorphisms with oral squamous cell carcinoma in a Northeastern Brazilian population: A pilot study

Objective to evaluate the association between XPD and XRCC3 polymorphisms and oral squamous cell carcinoma (OSCC). Design the sample consisted of 54 cases of OSCC and 40 cases of inflammatory fibrous hyperplasia (IFH). Genotypes were determined by the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. Results XPD-Lys/Gln was more common in IFH (n = 28; 70%) than in OSCC (n = 24; 44.4%) (OR: 0.3; p < 0.05). XPD-Gln was more frequent in high-grade lesions (0.48) than in low-grade lesions (0.21) (OR: 3.4; p < 0.05). The Gln/Gln genotype was associated with III and IV clinical stages (OR: 0.07; p < 0.05). XRCC3-Met was more frequent in OSCC (0.49) than in IFH (0.35) (OR: 2.6; p < 0.05). The Met/Met genotype was associated with the presence of metastases (OR: 8.1; p < 0.05) and with III and IV clinical stages (OR: 0.07; p < 0.05). Conclusions in this sample, the frequency of XPD-Gln in IFH suggests that this variant may protect against OSCC. The presence of the XRCC3-Met allele seems to contribute to the development of OSCC, metastases and more advanced stages in these lesions.

Papel da proteína de reparo XPC na regulação das proteínas de reparo APE1, OGG1 e PARP-1 em células humanas e de camundongos

studies using UV as a source of DNA damage. However, even though unrepaired UV-induced DNA damages are related to mutagenesis, cell death and tumorigenesis, they do not explain phenotypes such as neurodegeneration and internal tumors observed in patients with syndromes like Xeroderma Pigmentosum (XP) and Cockayne Syndrome (CS) that are associated with NER deficiency. Recent evidences point to a role of NER in the repair of 8-oxodG, a typical substrate of Base Excision Repair (BER). Since deficiencies in BER result in genomic instability, neurodegenerative diseases and cancer, it was investigated in this research the impact of XPC deficiency on BER functions in human cells. It was analyzed both the expression and the cellular localization of APE1, OGG1 e PARP-1, the mainly BER enzymes, in different NER-deficient human fibroblasts. The endogenous levels of these enzymes are reduced in XPC deficient cells. Surprisingly, XP-C fibroblasts were more resistant to oxidative agents than the other NER deficient fibroblasts, despite presenting the highest of 8-oxodG. Furthermore, subtle changes in the nuclear and mitochondrial localization of APE1 were detected in XP-C fibroblasts. To confirm the impact of XPC deficiency in the regulation of APE1 and OGG1 expression and activity, we constructed a XPC-complemented cell line. Although the XPC complementation was only partial, we found that XPC-complemented cells presented increased levels of OGG1 than XPC-deficient cells. The extracts from XPC-complemented cells also presented an elevated OGG1 enzimatic activity. However, it was not observed changes in APE1 expression and activity in the XPCcomplemented cells. In addition, we found that full-length APE1 (37 kDa) and OGG1- α are in the mitochondria of XPC-deficient fibroblasts and XPC-complemented fibroblasts before and after induction of oxidative stress. On the other hand, the expression of APE1 and PARP-1 are not altered in brain and liver of XPC knockout mice. However, XPC deficiency changed the APE1 localization in hypoccampus and hypothalamus. We also observed a physical interaction between XPC and APE1 proteins in human cells. In conclusion, the data suggest that XPC protein has a role in the regulation of OGG1 expression and activity in human cells and is involved mainly in the regulation of APE1 localization in mice. Aditionally, the response of NER deficient cells under oxidative stress may not be only associated to the NER deficiency per se, but it may include the new functions of NER enzymes in regulation of expression and cell localization of BER proteins