OBJECTIVE: To study the effect of hepatitis C virus nonstructural protein NS(3) (HCV NS3) on telomerase activity and carcinogenesis. METHODS: Streptavidin-peroxidase (SP) conjugated method was used to detect the expre...OBJECTIVE: To study the effect of hepatitis C virus nonstructural protein NS(3) (HCV NS3) on telomerase activity and carcinogenesis. METHODS: Streptavidin-peroxidase (SP) conjugated method was used to detect the expression of HCV NS(3) protein in NIH3T3 cells transfected with plasmid pRcHCNS(3)-5' and pRcHCNS(3)-3'. Telomerase activity was detected by an in situ telomerase activity labeling method, telomeric repeat amplification protocol polymerase chain reaction (TRAP-PCR) and telomerase PCR enzyme linked immunosorbent assay (ELISA) technology in the transfected and non-transfected NIH3T3 cells. RESULTS: HCV NS(3) protein was expressed in the NIH3T3 cells transfected with plasmid pRcHCNS(3)-5' expressing HCV NS(3) C-terminal deleted protein or with plasmid pRcHCNS(3)-3' expressing HCV NS(3) N-terminal deleted protein. The positive signal of HCV NS(3) protein was localized in the cytoplasm of NIH3T3 cells, and the signal intensity of the former was stronger. Telomerase activity in NIH3T3 cells transfected with plasmid pRcHCNS(3)-5' was stronger than that in NIH3T3 cells transfected with plasmid pRcHCNS(3)-3' (P展开更多
To study the effect of hepatitis C virus nonstructural protein NS 3 (HCV NS3) on telomerase activity and carcinogenesis Methods Streptavidin peroxidase (SP) conjugated method was used to detect the expressio n of...To study the effect of hepatitis C virus nonstructural protein NS 3 (HCV NS3) on telomerase activity and carcinogenesis Methods Streptavidin peroxidase (SP) conjugated method was used to detect the expressio n of HCV NS 3 protein in NIH3T3 cells transfected with plasmid pRcHCNS 3 5’ and pRcHCNS 3 3’ Telomerase activity was detected by an in situ telomerase a ctivity labeling method, telomeric repeat amplification protocol polymerase chai n reaction (TRAP PCR) and telomerase PCR enzyme linked immunosorbent assay (ELI SA) technology in the transfected and non transfected NIH3T3 cells Results HCV NS 3 protein was expressed in the NIH3T3 cells transfected with plasmid pR cHCNS 3 5’ expressing HCV NS 3 C terminal deleted protein or with plasmid pR cHCNS 3 3’ expressing HCV NS 3 N terminal deleted protein The positive sig nal of HCV NS 3 protein was localized in the cytoplasm of NIH3T3 cells, and th e signal intensity of the former was stronger Telomerase activity in NIH3T3 c ells transfected with plasmid pRcHCNS 3 5’ was stronger than that in NIH3T3 c ells transfected with plasmid pRcHCNS 3 3’ ( P 【0 01), whereas telomerase a ctivity in NIH3T3 cells transfected with plasmid pRcCMV or untreated NIH3T3 ce lls was weaker than that in NIH3T3 cells transfected with plasmid pRcHCNS 3 3 ’ ( P 【0 05) The expression level of HCV NS 3 protein was significantly co rrelated with the strength of telomerase activity ( P 【0 05) The results ob tained by in situ telomerase activity labeling corresponded to the results by te lomerase PCR ELISA technology Conclusions HCV NS 3 protein may activate telomerase through endogenous mechanism to induce host cell transformation The effect of HCV NS 3 C terminal deleted protein on telomerase activity in the host cell may be stronger than that of HCV NS 3 N terminal deleted protein In situ telomerase activity labeling was a reliabl e technology for studying pathological morphology and telomerase activity in tis sues and cells展开更多
Triple-negative breast cancer is an aggressive subtype that frequently develops resistance to chemotherapy. It is expected to develop new anti-tumor drugs through targeting the structure of G-quadruplexes of the genes...Triple-negative breast cancer is an aggressive subtype that frequently develops resistance to chemotherapy. It is expected to develop new anti-tumor drugs through targeting the structure of G-quadruplexes of the genes associated with this tumor. In this work, by targeting the 21-mer telomere G-quadruplex structure, compounds VB07 and VC02 were identified to stabilize the telomere G-quadruplex through structure-based high-throughput virtual screening. Cell cytotoxicity assay showed that VB07 and VC02 exhibited inhibitory effect on triple-negative breast cancer cells at the concentration of 5 μM. This study showed that structure-based high-throughput virtual screening was able to successfully identify the proper compounds targeting the telomere G-quadruplex, which exhibited inhibitory effects against the triple-negative breast cancer cells.展开更多
文摘OBJECTIVE: To study the effect of hepatitis C virus nonstructural protein NS(3) (HCV NS3) on telomerase activity and carcinogenesis. METHODS: Streptavidin-peroxidase (SP) conjugated method was used to detect the expression of HCV NS(3) protein in NIH3T3 cells transfected with plasmid pRcHCNS(3)-5' and pRcHCNS(3)-3'. Telomerase activity was detected by an in situ telomerase activity labeling method, telomeric repeat amplification protocol polymerase chain reaction (TRAP-PCR) and telomerase PCR enzyme linked immunosorbent assay (ELISA) technology in the transfected and non-transfected NIH3T3 cells. RESULTS: HCV NS(3) protein was expressed in the NIH3T3 cells transfected with plasmid pRcHCNS(3)-5' expressing HCV NS(3) C-terminal deleted protein or with plasmid pRcHCNS(3)-3' expressing HCV NS(3) N-terminal deleted protein. The positive signal of HCV NS(3) protein was localized in the cytoplasm of NIH3T3 cells, and the signal intensity of the former was stronger. Telomerase activity in NIH3T3 cells transfected with plasmid pRcHCNS(3)-5' was stronger than that in NIH3T3 cells transfected with plasmid pRcHCNS(3)-3' (P
文摘To study the effect of hepatitis C virus nonstructural protein NS 3 (HCV NS3) on telomerase activity and carcinogenesis Methods Streptavidin peroxidase (SP) conjugated method was used to detect the expressio n of HCV NS 3 protein in NIH3T3 cells transfected with plasmid pRcHCNS 3 5’ and pRcHCNS 3 3’ Telomerase activity was detected by an in situ telomerase a ctivity labeling method, telomeric repeat amplification protocol polymerase chai n reaction (TRAP PCR) and telomerase PCR enzyme linked immunosorbent assay (ELI SA) technology in the transfected and non transfected NIH3T3 cells Results HCV NS 3 protein was expressed in the NIH3T3 cells transfected with plasmid pR cHCNS 3 5’ expressing HCV NS 3 C terminal deleted protein or with plasmid pR cHCNS 3 3’ expressing HCV NS 3 N terminal deleted protein The positive sig nal of HCV NS 3 protein was localized in the cytoplasm of NIH3T3 cells, and th e signal intensity of the former was stronger Telomerase activity in NIH3T3 c ells transfected with plasmid pRcHCNS 3 5’ was stronger than that in NIH3T3 c ells transfected with plasmid pRcHCNS 3 3’ ( P 【0 01), whereas telomerase a ctivity in NIH3T3 cells transfected with plasmid pRcCMV or untreated NIH3T3 ce lls was weaker than that in NIH3T3 cells transfected with plasmid pRcHCNS 3 3 ’ ( P 【0 05) The expression level of HCV NS 3 protein was significantly co rrelated with the strength of telomerase activity ( P 【0 05) The results ob tained by in situ telomerase activity labeling corresponded to the results by te lomerase PCR ELISA technology Conclusions HCV NS 3 protein may activate telomerase through endogenous mechanism to induce host cell transformation The effect of HCV NS 3 C terminal deleted protein on telomerase activity in the host cell may be stronger than that of HCV NS 3 N terminal deleted protein In situ telomerase activity labeling was a reliabl e technology for studying pathological morphology and telomerase activity in tis sues and cells
基金National Natural Science Foundation of China(Grant No.31701791,21732002,31672558 and 21502060)Huazhong Agricultural University Scientific&Technological Self-innovation Foundation(Grant No.2662017PY113,2015RC013 and 2662015PY208)Open fund of The State Key Laboratory of Bio-organic and Natural Products Chemistry,CAS(Grant No.SKLBNPC16343)。
文摘Triple-negative breast cancer is an aggressive subtype that frequently develops resistance to chemotherapy. It is expected to develop new anti-tumor drugs through targeting the structure of G-quadruplexes of the genes associated with this tumor. In this work, by targeting the 21-mer telomere G-quadruplex structure, compounds VB07 and VC02 were identified to stabilize the telomere G-quadruplex through structure-based high-throughput virtual screening. Cell cytotoxicity assay showed that VB07 and VC02 exhibited inhibitory effect on triple-negative breast cancer cells at the concentration of 5 μM. This study showed that structure-based high-throughput virtual screening was able to successfully identify the proper compounds targeting the telomere G-quadruplex, which exhibited inhibitory effects against the triple-negative breast cancer cells.
