Light is an environmental signaling,whereas Aux/IAA proteins and Auxin Response Factors(ARFs)are regulators of auxin signalling.Aux/IAA proteins are unstable,and their degradation dependents on 26S ubiquitin-proteasom...Light is an environmental signaling,whereas Aux/IAA proteins and Auxin Response Factors(ARFs)are regulators of auxin signalling.Aux/IAA proteins are unstable,and their degradation dependents on 26S ubiquitin-proteasome and is promoted by Auxin.Auxin binds directly to a SCF-type ubiquitin-protein ligase,TIR1,facilitates the interaction between Aux/IAA proteins and TIR1,and then the degradation of Aux/IAA proteins.A few studies have reported that some ARFs are also unstable proteins,and their degradation is also mediated by 26S proteasome.In this study,by using of antibodies recognizing endogenous ARF7 proteins,we found that protein stability of ARF7 was affected by light.By expressing MYC tagged ARF activators in protoplasts,we found that degradation of ARF7 was inhibited by 26 proteasome inhibitors.In addition,at least ARF5 and ARF19 were also unstable proteins,and degradation of ARF5 via 26S proteasome was further confirmed by using stable transformed plants overexpressing ARF5 with a GUS tag.展开更多
Objective To investigate the synergistic antitumor effects of combined use of p14ARF gene and 5-fluorouracil (5-Fu) in pancreatic cancer.Methods A human pancreatic cancer cell line PC-3 was transfected with lipofect...Objective To investigate the synergistic antitumor effects of combined use of p14ARF gene and 5-fluorouracil (5-Fu) in pancreatic cancer.Methods A human pancreatic cancer cell line PC-3 was transfected with lipofectin-mediated recombinant p14ARF gene,and was then administered with 5-Fu. Cell growth,morphological changes,cell cycle,apoptosis,and molecular changes were measured using the MTT assay,flow cytometry,RT-PCR,Western blotting,and immunocytochemical assays.Results After transfection of p14ARF,cell growth was obviously inhibited,resulting in an accumulation of cells in the G 1 phase. The proportion of cells in the G 1 phase was significantly increased from 58.51% to 75.92 %,and in the S and G 2/M phases decreased significantly from 20.05% to 12.60%,and from 21.44% to 11.48 %,respectively,as compared with those of the control groups. PC-3/p14ARF cells that underwent 5-Fu treatment had significantly greater G 2/M phase accumulation,from 11.48% to 53.47 %. The apoptopic index was increased in PC-3/p14ARF cells from 3.64% to 19.62%. The MTT assay showed p14ARF-expressing cells were significantly more sensitive to 5-Fu (0.01-10 mg/L) than those devoid of p14ARF expression ( P <0.01). Western blotting showed p14ARF upregulates p53 expression. Conclusion Combined use of p14ARF gene and 5-Fu acts synergistically to inhibit pancreatic cancer cell proliferation,suggesting a new anticancer strategy.展开更多
文摘Light is an environmental signaling,whereas Aux/IAA proteins and Auxin Response Factors(ARFs)are regulators of auxin signalling.Aux/IAA proteins are unstable,and their degradation dependents on 26S ubiquitin-proteasome and is promoted by Auxin.Auxin binds directly to a SCF-type ubiquitin-protein ligase,TIR1,facilitates the interaction between Aux/IAA proteins and TIR1,and then the degradation of Aux/IAA proteins.A few studies have reported that some ARFs are also unstable proteins,and their degradation is also mediated by 26S proteasome.In this study,by using of antibodies recognizing endogenous ARF7 proteins,we found that protein stability of ARF7 was affected by light.By expressing MYC tagged ARF activators in protoplasts,we found that degradation of ARF7 was inhibited by 26 proteasome inhibitors.In addition,at least ARF5 and ARF19 were also unstable proteins,and degradation of ARF5 via 26S proteasome was further confirmed by using stable transformed plants overexpressing ARF5 with a GUS tag.
基金ThisinvestigationwaspartlysupportedbyagrantfromtheShanghaiScienceFundation (No 0 0 41190 0 9)
文摘Objective To investigate the synergistic antitumor effects of combined use of p14ARF gene and 5-fluorouracil (5-Fu) in pancreatic cancer.Methods A human pancreatic cancer cell line PC-3 was transfected with lipofectin-mediated recombinant p14ARF gene,and was then administered with 5-Fu. Cell growth,morphological changes,cell cycle,apoptosis,and molecular changes were measured using the MTT assay,flow cytometry,RT-PCR,Western blotting,and immunocytochemical assays.Results After transfection of p14ARF,cell growth was obviously inhibited,resulting in an accumulation of cells in the G 1 phase. The proportion of cells in the G 1 phase was significantly increased from 58.51% to 75.92 %,and in the S and G 2/M phases decreased significantly from 20.05% to 12.60%,and from 21.44% to 11.48 %,respectively,as compared with those of the control groups. PC-3/p14ARF cells that underwent 5-Fu treatment had significantly greater G 2/M phase accumulation,from 11.48% to 53.47 %. The apoptopic index was increased in PC-3/p14ARF cells from 3.64% to 19.62%. The MTT assay showed p14ARF-expressing cells were significantly more sensitive to 5-Fu (0.01-10 mg/L) than those devoid of p14ARF expression ( P <0.01). Western blotting showed p14ARF upregulates p53 expression. Conclusion Combined use of p14ARF gene and 5-Fu acts synergistically to inhibit pancreatic cancer cell proliferation,suggesting a new anticancer strategy.
