RESUMO
OBJECTIVE: To analyze how the infection of human papillomavirus 16 (HPV16) affects expression of DNA polymerase beta (DNA polB) with the aim of probing the mechanism of over-expression of DNA polB in human cancers. METHODS: Four fragments of human DNA polB promoter were amplified and constructed into luciferase reporter vector pGL-Basic, generating pGL-BP, pGL-BMH, pGL-BMS, and pGL-BAT constructs respectively, and co-transfected with HPV16 or HPV6 into Hep2 cells. Luciferase activity was assayed 48 hours after transfection. Semi-quantitative RT-PCR was used to measure mRNA expression of endogenous DNA polB. Immunohistochemistry and in situ hybridization were used to analyze DNA polB expression and HPV16 or HPV6 infection in 38 cases of cervical lesions respectively. RESULTS: With co-transfection of HPV16 and DNA polB promoter-driving reporters into Hep2 cells, pGL-BP reporter in full-length DNA polB promoter presented markedly elevated luciferase activities (P < 0.05). However, the other three mutant reporters: pGL-BMH, pGL-BMS, and pGL-BAT, generated no reporting activities in the presence of HPV16 (P > 0.05). On the contrary, all of polB promoter reporters were little stimulated in co-transfection of HPV6 (P > 0.05). The transfection of HPV16 could enhance the endogenous polB mRNA expression compared with that of HPV6 (3.42 vs. 0.80, P < 0.05). The DNA polB expression was found in 8 of 10 HPV16-positive cervical intraepithelial neoplasia grade III (CIN III) cases, while was only found in 3 of 11 HPV6-positive condyloma accuminatum cases, but was negative in all chronic cervicitis cases. The correlation of DNA polB expression with HPV16 infection in cervical lesions was significant (P < 0.05). CONCLUSIONS: HPV16 is able to specifically stimulate the expression of DNA polB in human epithelial cells through interaction with the core upstream regulatory sequences of DNA polB promoter. Over-expression of DNA polB might be an explanation for the molecular mechanism underlying HPV-related human cancers.
Assuntos
DNA Polimerase beta/metabolismo , Regulação Enzimológica da Expressão Gênica , Papillomavirus Humano 16/metabolismo , Linhagem Celular , DNA Polimerase beta/genética , Feminino , Papillomavirus Humano 16/genética , Papillomavirus Humano 6/genética , Papillomavirus Humano 6/metabolismo , Humanos , Infecções por Papillomavirus , Regiões Promotoras Genéticas , Neoplasias do Colo do Útero/genética , Neoplasias do Colo do Útero/patologia , Neoplasias do Colo do Útero/virologiaRESUMO
A taxonomic study was performed on strain QM42(T), which was isolated from coastal seawater from an aquaculture site near Qingdao, China. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain QM42(T) was a member of the class Gammaproteobacteria. Cells of strain QM42(T) were Gram-negative, yellow, aerobic and rod-shaped. The strain formed a distinct phyletic line with less than 91 % 16S rRNA gene sequence similarity to its closest relatives with validly published names within the class Gammaproteobacteria. The genomic DNA G+C content was 51.9 mol%. The major fatty acids were C(16 : 1)omega7c/iso-C(15 : 0) 2-OH, C(18 : 1)omega7c and C(16 : 0). Based on data from a polyphasic chemotaxonomic, physiological and biochemical study, strain QM42(T) is considered to represent a novel genus and species, for which the name Gilvimarinus chinensis gen. nov., sp. nov., is proposed. The type strain is QM42(T) (=CGMCC 1.7008(T)=DSM 19667(T)).