The partitioning-defective 3 （Par3）, a key component in the conserved Par3/Par6/aPKC complex, plays fundamental roles in cell polarity. Herein we report the identification of Ku70 and Ku80 as novel Par3-interacting ...The partitioning-defective 3 （Par3）, a key component in the conserved Par3/Par6/aPKC complex, plays fundamental roles in cell polarity. Herein we report the identification of Ku70 and Ku80 as novel Par3-interacting proteins through an in vitro binding assay followed by liquid chromatography-tandem mass spectrometry. Ku70/Ku80 proteins are two key regulatory subunits of the DNA-dependent protein kinase （DNA-PK）, which plays an essential role in repairing double-strand DNA breaks （DSBs）. We determined that the nuclear association of Par3 with Ku70/Ku80 was enhanced by γ-irradiation （IR）, a potent DSB inducer. Furthermore, DNA-PKcs, the catalytic subunit of DNA-PK, interacted with the Par3/Ku70/Ku80 complex in response to IR. Par3 over-expression or knockdown was capable of up- or downregulating DNA-PK activity, respectively. Moreover, the Par3 knockdown cells were found to be defective in random plasmid integration, defective in DSB repair following IR, and radiosensitive, phenotypes similar to that of Ku70 knockdown cells. These findings identify Par3 as a novel component of the DNA-PK complex and implicate an unexpected link of cell polarity to DSB repair.展开更多
The author affiliations were mixed up in the previous published version. The third fund number of National NaturalScience Foundation of China in the Acknowledgments was wrong, it should be "30270335". The Sh...The author affiliations were mixed up in the previous published version. The third fund number of National NaturalScience Foundation of China in the Acknowledgments was wrong, it should be "30270335". The Shanghai MunicipalCouncil for Science and Technology (No.06DZ22032) was missed in the Acknowledgments. There are some labelingand production errors in Figure 2A, Figure 3B and 3C, Figure 5C, Figure 6B and 6E, Figure 7B and 7D. In Figure 2A,left panel "A431" should be "Par3". In Figure 3B and 3C, "anti-Par3CT" should be "anti-Par3LCT", "GST-Par3CT"should be "GST-Par3LCT". In Figure 5C, the second arrow indicating "Lamin B" should be "[3-tubulin". In Figure 6Bright panel, the molecular weight for ?-actin should be "43" instead of"200". In Figure 6E, "Par3" should be "Par3i". Themolecular weight for the DNA-PKcs panel should be the same as the p-DNA-PKcs. In Figure 7B, the time point "240"in the left panel should be "120"; in the right panel of Figure 7B, the title for the y axis should be "DNA released (%)".In Figure 7D, the title for the y axis should be "Survival (%)", and the scale for the y axis should be "100, 10 and 1". These corrections do not affect the conclusions of the study. We apologize for any inconvenience this may havecaused.展开更多
Biophotonics is an exciting and fast-expanding frontier which involves the fusion of advanced photonics and biology.It has not only created many novel methodologies for biomedical research,but also achieved many signi...Biophotonics is an exciting and fast-expanding frontier which involves the fusion of advanced photonics and biology.It has not only created many novel methodologies for biomedical research,but also achieved many significant results as an independentfield.Thanks to femtosecond(fs)laser technologies,important progresses have been made regarding the manipulation,imaging,and engineering of biological samples ranging from single molecules to tissues in the last 20 years.The ultrashort pulses at near-infrared band provide many advantages:high nonlinear efficiency,low absorption by biological samples,high spatial and temporal resolution and confinement,and low phototoxicity.They are noninvasive and easy to control.Although the mechanism of how fs laser pulses interact with cells remains unclear,experimental results have shown that they could open up the cell membrane and hence made optical transfection and optical cell fusion possible.In this review,some of the seminal works on transfection and cell fusion by fs lasers are presented.The ideas behind and the experimental details will be described together with a highlight on their significances.Specifically,the thermal effect is analyzed based on multiphoton excitation and plasma formation in an aqueous environment to explain the nontoxic characteristic of fs laser irradiation.Last,some applications of fs laser induced transfection and cellcell fusion with potential major impact in biomedical sciences are proposed.展开更多
基金This work was supported by the grants from National Natural Science Foundation of China(Nos.30170208,30623003 and 30170208)from the Ministry of Science and Technology,China(No.2001AA233031 and No.2001CB510205)from US NIH(CA50519)(to DJ Chen).
文摘The partitioning-defective 3 （Par3）, a key component in the conserved Par3/Par6/aPKC complex, plays fundamental roles in cell polarity. Herein we report the identification of Ku70 and Ku80 as novel Par3-interacting proteins through an in vitro binding assay followed by liquid chromatography-tandem mass spectrometry. Ku70/Ku80 proteins are two key regulatory subunits of the DNA-dependent protein kinase （DNA-PK）, which plays an essential role in repairing double-strand DNA breaks （DSBs）. We determined that the nuclear association of Par3 with Ku70/Ku80 was enhanced by γ-irradiation （IR）, a potent DSB inducer. Furthermore, DNA-PKcs, the catalytic subunit of DNA-PK, interacted with the Par3/Ku70/Ku80 complex in response to IR. Par3 over-expression or knockdown was capable of up- or downregulating DNA-PK activity, respectively. Moreover, the Par3 knockdown cells were found to be defective in random plasmid integration, defective in DSB repair following IR, and radiosensitive, phenotypes similar to that of Ku70 knockdown cells. These findings identify Par3 as a novel component of the DNA-PK complex and implicate an unexpected link of cell polarity to DSB repair.
基金supported by the grants from National Natural Science Foundation of China(No.30170208,30623003 and 30270335)from the Ministry of Science and Technology,China(No.2001AA233031 and No.2001CB510205)+1 种基金from the Shanghai Municipal Council for Science and Technology(No.06DZ22032)from US NIH(CA50519)(to DJ Chen).
文摘The author affiliations were mixed up in the previous published version. The third fund number of National NaturalScience Foundation of China in the Acknowledgments was wrong, it should be "30270335". The Shanghai MunicipalCouncil for Science and Technology (No.06DZ22032) was missed in the Acknowledgments. There are some labelingand production errors in Figure 2A, Figure 3B and 3C, Figure 5C, Figure 6B and 6E, Figure 7B and 7D. In Figure 2A,left panel "A431" should be "Par3". In Figure 3B and 3C, "anti-Par3CT" should be "anti-Par3LCT", "GST-Par3CT"should be "GST-Par3LCT". In Figure 5C, the second arrow indicating "Lamin B" should be "[3-tubulin". In Figure 6Bright panel, the molecular weight for ?-actin should be "43" instead of"200". In Figure 6E, "Par3" should be "Par3i". Themolecular weight for the DNA-PKcs panel should be the same as the p-DNA-PKcs. In Figure 7B, the time point "240"in the left panel should be "120"; in the right panel of Figure 7B, the title for the y axis should be "DNA released (%)".In Figure 7D, the title for the y axis should be "Survival (%)", and the scale for the y axis should be "100, 10 and 1". These corrections do not affect the conclusions of the study. We apologize for any inconvenience this may havecaused.
基金This work was supported in part by The Research Grants Council of HKSAR Government under GRF Grant Nos.CUHK410708 and 410809.
文摘Biophotonics is an exciting and fast-expanding frontier which involves the fusion of advanced photonics and biology.It has not only created many novel methodologies for biomedical research,but also achieved many significant results as an independentfield.Thanks to femtosecond(fs)laser technologies,important progresses have been made regarding the manipulation,imaging,and engineering of biological samples ranging from single molecules to tissues in the last 20 years.The ultrashort pulses at near-infrared band provide many advantages:high nonlinear efficiency,low absorption by biological samples,high spatial and temporal resolution and confinement,and low phototoxicity.They are noninvasive and easy to control.Although the mechanism of how fs laser pulses interact with cells remains unclear,experimental results have shown that they could open up the cell membrane and hence made optical transfection and optical cell fusion possible.In this review,some of the seminal works on transfection and cell fusion by fs lasers are presented.The ideas behind and the experimental details will be described together with a highlight on their significances.Specifically,the thermal effect is analyzed based on multiphoton excitation and plasma formation in an aqueous environment to explain the nontoxic characteristic of fs laser irradiation.Last,some applications of fs laser induced transfection and cellcell fusion with potential major impact in biomedical sciences are proposed.
