RESUMO
Persistent infections by high-risk human papillomavirus (HR-HPV) are a necessary condition, but not sufficient for cervical cancer development. Genetic variants of HR-HPV appear to be related to the risk of persistent infections. The study performed a functional evaluation of variants of the HPV-31 promoter region (LCR). For this, cloning and subcloning of variants HPV-31/UFPE-21 HPV-31/UFPE-89, HPV-31/UFPE-66, E2 gene and prototype HPV-31 were performed. Transfection with different concentrations of E2 was done and the concentration of 25 ng was determined to be ideal for LCR activation. HPV-31/UFPE-21 and HPV-31/UFPE-89 have a greater ability to alter Nluc reporter gene expression levels and HPV-31/UFPE-66 showed decreased levels of gene expression of Nluc reporter gene compared to control. Statistical analysis showed a significant difference between the polymorphic LCR regions and the control (p < 0.0001). A more refined profile of variants of HPV-31 and its importance for the prognosis of cervical lesions begins to be drawn.
Assuntos
Papillomavirus Humano 31/genética , Regiões Promotoras Genéticas , Proteínas de Ligação a DNA/metabolismo , Células HeLa , Humanos , Polimorfismo Genético , Transativadores/metabolismo , Ativação Transcricional , Proteínas Virais/metabolismoRESUMO
Persistent infections caused by high-risk human papillomavirus (HR-HPV) are important, for the development of cervical lesions, but environmental and genetic factors are also related in the process of carcinogenesis. Among the genetic factors, the genetic variants of HR-HPV appear to be related to the risk of persistent infections. Therefore, the present study investigates variants of HPV31 E5 oncogene in cervical scraping samples from Brazilian women to assess their functional and structural effects, in order to identify possible repercussions of these variants on the infectious and carcinogenic process. Our results detected nucleotide changes previously described in the HPV31 E5 oncogene, which may play a critical role in the development of cancer due to its ability to promote cell proliferation and signal transmission. In our study, the interaction percentage of the 31E5 sequence generated by the Immune Epitope Server database and the Analysis Resource (IEDB) allowed us to include possible immunogenic epitopes with the MHC-I and MHC-II molecules, which may represent a possible relationship between protein suppression of the immune system. In the structural analysis of the HPV31 E5 oncoprotein, the N5D, I48 V, P56A, F80I and V64I polymorphisms can be found inserted within transmembrane regions. The P56A mutation has been predicted to be highly stabilizing and, therefore, can cause a change in protein function. Regarding the interaction of the E5 protein from HPV31 with the signaling of NF-kB pathway, we observed that in all variants of the E5 gene from HPV-31, the activity of the NF-kB pathway was increased compared to the prototype. Our study contributes to a more refined design of studies with the E5 gene from HPV31 and provides important data for a better understanding of how variants can be distinguished under their clinical consequences.
Assuntos
Colo do Útero/virologia , Variação Genética , Papillomavirus Humano 31/classificação , Papillomavirus Humano 31/genética , Proteínas Oncogênicas Virais/química , Proteínas Oncogênicas Virais/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Proliferação de Células , Feminino , Células HEK293 , Humanos , Pessoa de Meia-Idade , Mutação , Proteínas Oncogênicas Virais/classificação , Infecções por Papillomavirus/virologia , Filogenia , Proteínas Serina-Treonina Quinases/genética , Transdução de Sinais , Neoplasias do Colo do Útero/virologia , Adulto Jovem , Quinase Induzida por NF-kappaBRESUMO
BACKGROUND: The MDM2 gene is the major negative regulator of p53, a tumor suppressor protein. Single nucleotide polymorphism in promoter region of MDM2 gene leads to increased expression resulting in higher levels of MDM2 protein. This event increases the attenuation of the p53 pathway. Polymorphisms in this gene can interfere in the regulation of cellular proliferation. We evaluated whether MDM2 SNP309 (rs2278744) associated or not with the use of oral contraceptive can heighten susceptibility to development of cervical lesions in women HPV infected. METHODS: MDM2 SNP309 (rs2278744) was genotyped in a total of 287 patients using the PCR-RFLP technique. The results were analyzed by UNPHASED v.3.121 and SNPStats programs. RESULTS: The three groups (SIL, LSIL and HSIL) showed no significant differences in either genotype or allelic frequencies for MDM2 polymorphisms, except when HSIL was compared with LSIL (p = 0.037; OR = 1.81). Furthermore, in the analysis of contraceptives, a significant association was found between the use of contraceptives and the MDM2 variant in the development of high-grade cervical lesions for the TG genotype (p = 0.019; OR = 2.21) when HSIL was compared with control. When HSIL was compared with LSIL (p = 0.006; OR = 2.27). CONCLUSION: The results of this study suggest that MDM2 SNP309 might be a good marker for assessing the progression of LSIL to HSIL. In addition, they also show that oral contraceptives alone, did not have any effect on the progression or development of cervical lesions. However, they may act synergistically with MDM2 SNP309 (rs2278744) and HPV infection in the development of cervical lesions.
RESUMO
OBJECTIVE: We sought to characterize E6 and E7 oncogenes genetic variability of HPV-31 isolated from cervical scraping samples of Northeastern Brazilian women. METHODS: E6 and E7 were amplified with specific primers, cloned and sequenced. The sequences obtained were aligned with the GenBank reference sequences with the aim of evaluating the possible genetic variants. RESULTS: We identified several genetic variants in E6 and E7 sequences from HPV-31 positive women. Three nucleotide changes in E6 were described for the first time in this study. Some nucleotide changes were non-synonymous substitutions. CONCLUSION: The knowledge of region/country HPV specific genetic variations is relevant to understand the epidemiology and the development of effective vaccines.