RESUMO
Chronic infections with the hepatitis B virus (HBV) and high-risk human papillomaviruses (HPVs) are important risk factors for hepatocellular carcinoma (HCC) and cervical cancer (CC), respectively. HBV and HPV are DNA viruses that almost invariably integrate into the host genome in invasive tumors. The viral integration sites occur throughout the genome, leading to the presumption that there are no preferred sites of integration. A number of viral integrations have been shown to occur within the vicinity of important cancer-related genes. In studies of HBV-induced HCC and HPV-induced CC, we have identified two HBV and three HPV integrations into the human telomerase reverse transcriptase (hTERT) gene. Detailed characterization of the integrations revealed that four integrations occurred within the hTERT promoter and upstream region and the fifth integration occurred in intron 3 of the hTERT gene. None of the integrations altered the hTERT coding sequence and all resulted in juxtaposition of viral enhancers near hTERT, with potential activation of hTERT expression. Our work supports the hypothesis that the sites of oncogenic viral integration are nonrandom and that genes at the sites of viral integration may play important roles in carcinogenesis.
Assuntos
Vírus da Hepatite B/genética , Neoplasias Hepáticas/virologia , Papillomaviridae/genética , Telomerase/genética , Neoplasias do Colo do Útero/virologia , Integração Viral , Sequência de Bases , Proteínas de Ligação a DNA , Feminino , Regulação Enzimológica da Expressão Gênica , Humanos , Neoplasias Hepáticas/enzimologia , Neoplasias Hepáticas/etiologia , Dados de Sequência Molecular , Células Tumorais Cultivadas , Neoplasias do Colo do Útero/enzimologia , Neoplasias do Colo do Útero/etiologiaRESUMO
The role of microsatellite instability (MSI) in the pathogenesis of hepatocellular carcinoma (HCC) is incompletely defined. Although high-frequency MSI (MSI-H) is infrequently seen in HCC, some studies have suggested a role for MSI in HCC development. While MSI has been clearly defined for a subset of tumors, in particular colorectal, gastric and endometrial cancers, generally accepted criteria have not been developed for other tumors. Colorectal cancers (CRC) are classified as MSI-H if >30-40% of >5 microsatellite loci analyzed show instability. The MSI-H phenotype is associated with defective DNA mismatch repair (MMR) and is observed in the majority of tumors from patients with hereditary non-polyposis colon cancer (HNPCC) and also in 15% of sporadic CRCs. Inactivating mutations of the hMLH1 or hMSH2 genes lead to defects in MMR in HNPCC. In sporadic CRCs, MMR is usually due to hypermethylation of the hMLH-1 promoter. The role of defective MMR in hepatocellular carcinogenesis is controversial. Immunohistochemistry for hMLH1 and hMSH2 reliably indicates hMLH1 or hMSH2 loss in MSI-H CRC tumors. To investigate the role of defective MMR in HCC carcinogenesis, we performed immunohistochemistry for hMLH1 and hMSH2 on 36 HCCs. BAT26, a microsatellite marker that reliably predicts MSI-H was also examined. All 36 of the tumors stained positively for both hMLH1 and hMSH2, strongly suggesting an absence of either inactivating mutations of hMLH1 and hMSH2 or promoter hypermethylation of hMLH1. None of the tumors showed MSI at the BAT26 locus. These findings suggest that defective MMR does not contribute significantly to hepatocellular carcinogenesis.
Assuntos
Pareamento Incorreto de Bases , Carcinoma Hepatocelular/metabolismo , Proteínas de Ligação a DNA , Neoplasias Hepáticas/metabolismo , Proteínas de Neoplasias/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Adaptadoras de Transdução de Sinal , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinoma Hepatocelular/genética , Proteínas de Transporte , Metilação de DNA , Primers do DNA/química , Feminino , Humanos , Técnicas Imunoenzimáticas , Neoplasias Hepáticas/genética , Masculino , Repetições de Microssatélites , Pessoa de Meia-Idade , Proteína 1 Homóloga a MutL , Proteína 2 Homóloga a MutS , Proteínas de Neoplasias/genética , Estadiamento de Neoplasias , Proteínas Nucleares , Reação em Cadeia da Polimerase , Proteínas Proto-Oncogênicas/genéticaRESUMO
The incorporation of 5-fluorouracil (FUra) into RNA of Mycobacterium avium complex strain LM1 was evaluated. Cells were labeled with either [14C]FUra or [3H]uracil and the ribonucleosides were analyzed by high-performance liquid chromatography. The identification of the ribonucleosides was facilitated by the use of an isocratic system that provided unambiguous identification of the RNA pyrimidine components. Uracil was incorporated into RNA as uridine, but an equal amount was converted to cytidine. [14C]FUra was incorporated directly into RNA as 5-fluorouridine and there was no evidence of its conversion to other pyrimidines. The ratio of 5-fluorouridine:uridine was 2.8-fold greater for cells grown in 100 micrograms FUra/mL than for cells grown in 20 micrograms FUra/mL. Analysis of the RNA nucleotides was performed and deoxyribonucleotides were present; DNA contamination was estimated to range from about 2 to 8% of the RNA preparations.
