The genomes of eukaryotic cells are under continuous assault by environmental agents and endogenous metabolic byproducts. Damage induced in DNA usually leads to a cascade of cellular events, the DNA damage response. F...The genomes of eukaryotic cells are under continuous assault by environmental agents and endogenous metabolic byproducts. Damage induced in DNA usually leads to a cascade of cellular events, the DNA damage response. Failure of the DNA damage response can lead to development of malignancy by reducing the efficiency and fidelity of DNA repair. The NBS1 protein is a component of the MRE11/RAD50/NBS 1 complex （MRN） that plays a critical role in the cellular response to DNA damage and the maintenance of chromosomal integrity. Mutations in the NBS1 gene are responsible for Nijmegen breakage syndrome （NBS）, a hereditary disorder that imparts an increased predisposition to development of malignancy. The phenotypic characteristics of cells isolated from NBS patients point to a deficiency in the repair of DNA double strand breaks. Here, we review the current knowledge of the role of NBS1 in the DNA damage response. Emphasis is placed on the role of NBS1 in the DNA double strand repair, modulation of the DNA damage sensing and signaling, cell cycle checkpoint control and maintenance oftelomere stability.展开更多
背景与目的:组蛋白去乙酰化酶1(sirtuin type 1,SIRT1)在胰腺癌等恶性肿瘤中高表达,可能与肿瘤的恶性表型相关。本研究采用RNA干扰技术下调胰腺癌PANC1细胞中SIRT1的表达,探讨其对DNA损伤修复及放化疗抵抗的影响。方法:用彗星试验检测...背景与目的:组蛋白去乙酰化酶1(sirtuin type 1,SIRT1)在胰腺癌等恶性肿瘤中高表达,可能与肿瘤的恶性表型相关。本研究采用RNA干扰技术下调胰腺癌PANC1细胞中SIRT1的表达,探讨其对DNA损伤修复及放化疗抵抗的影响。方法:用彗星试验检测细胞敲除SIRT1后DNA损伤修复能力;蛋白质印迹法(Westernblot)检测NBS1、γH2AX DNA修复相关蛋白的表达水平;最后用克隆存活实验和细胞增殖实验观察胰腺癌细胞对放化疗的敏感性。结果:下调SIRT1显著抑制了胰腺癌细胞的DNA损伤修复能力;抑制了DNA损伤修复相关蛋白NBS1磷酸化活化;增强了细胞对放化疗的敏感性。结论:SIRT1基因表达下调可抑制胰腺癌细胞DNA损伤修复能力,提高其对放化疗的敏感性,机制上可能与SIRT1下调后NBS1磷酸化活化受抑制相关。展开更多
Objective： To research the silence of NBS1 after transfection microRNA expressing eukaryotic recombinants and the changes of telomerase activation in telomerase-positive cell line Hela. Methods： According to the seq...Objective： To research the silence of NBS1 after transfection microRNA expressing eukaryotic recombinants and the changes of telomerase activation in telomerase-positive cell line Hela. Methods： According to the sequence of NBS1 mRNA, the NBS1 pre-microRNA was designed and synthesized, then cloned into the GFP reporter pcDNA6.2-GW/EmGFP-miR vector and transfected into Hela cells. The integrity of the insert fragment was verified through colony PCR and sequencing analysis. The NBS1 gene expression of NBS1 microRNA recombinants was detected by Real-Time PCR and western blot. Telomerase activity in Hela cells was assayed by TRAP-PCR-EB. Results： Sequences of insert fragment in microRNA expressing recombinants were correct. The NBS1 gene expression was decreased, and the telomerase activation of Hela cell reduced. Conclusion： NBS1 microRNA inhibits NBS1 gene expression, and depresses telomerase activation of Hela cells. This confirms that there is relevance between NBS 1 gene and telomerase activity.展开更多
Human single-stranded DNA-binding protein 1(hSSB1)is required for the efficient recruitment of the MRN complex to DNA doublestrand breaks and is essential for the maintenance of genome integrity.However,the mechanism ...Human single-stranded DNA-binding protein 1(hSSB1)is required for the efficient recruitment of the MRN complex to DNA doublestrand breaks and is essential for the maintenance of genome integrity.However,the mechanism by which hSSB1 recruits NBS1 remains elusive.Here,we determined that hSSB1 undergoes SUMOylation at both K79 and K94 under normal conditions and that this modification is dramatically enhanced in response to DNA damage.SUMOylation of hSSB1,which is specifically fine-tuned by PIAS2α,and SENP2,not only stabilizes the protein but also enhances the recruitment of NBS1 to DNA damage sites.Cells with defective hSSB1 SUMOylation are sensitive to ionizing radiation,and global inhibition of SUMOylation by either knocking out UBC9 or adding SUMOylation inhibitors significantly enhances the sensitivity of cancer cells to etoposide.Our findings reveal that SUMOylation,as a novel posttranslational modification of hSSB1,is critical for the functions of this protein,indicating that the use of SUMOylation inhibitors(e.g.,2-D08 and ML-792)may be a new strategy that would benefit cancer patients being treated with chemo-or radiotherapy.展开更多
文摘The genomes of eukaryotic cells are under continuous assault by environmental agents and endogenous metabolic byproducts. Damage induced in DNA usually leads to a cascade of cellular events, the DNA damage response. Failure of the DNA damage response can lead to development of malignancy by reducing the efficiency and fidelity of DNA repair. The NBS1 protein is a component of the MRE11/RAD50/NBS 1 complex （MRN） that plays a critical role in the cellular response to DNA damage and the maintenance of chromosomal integrity. Mutations in the NBS1 gene are responsible for Nijmegen breakage syndrome （NBS）, a hereditary disorder that imparts an increased predisposition to development of malignancy. The phenotypic characteristics of cells isolated from NBS patients point to a deficiency in the repair of DNA double strand breaks. Here, we review the current knowledge of the role of NBS1 in the DNA damage response. Emphasis is placed on the role of NBS1 in the DNA double strand repair, modulation of the DNA damage sensing and signaling, cell cycle checkpoint control and maintenance oftelomere stability.
文摘背景与目的:组蛋白去乙酰化酶1(sirtuin type 1,SIRT1)在胰腺癌等恶性肿瘤中高表达,可能与肿瘤的恶性表型相关。本研究采用RNA干扰技术下调胰腺癌PANC1细胞中SIRT1的表达,探讨其对DNA损伤修复及放化疗抵抗的影响。方法:用彗星试验检测细胞敲除SIRT1后DNA损伤修复能力;蛋白质印迹法(Westernblot)检测NBS1、γH2AX DNA修复相关蛋白的表达水平;最后用克隆存活实验和细胞增殖实验观察胰腺癌细胞对放化疗的敏感性。结果:下调SIRT1显著抑制了胰腺癌细胞的DNA损伤修复能力;抑制了DNA损伤修复相关蛋白NBS1磷酸化活化;增强了细胞对放化疗的敏感性。结论:SIRT1基因表达下调可抑制胰腺癌细胞DNA损伤修复能力,提高其对放化疗的敏感性,机制上可能与SIRT1下调后NBS1磷酸化活化受抑制相关。
文摘Objective： To research the silence of NBS1 after transfection microRNA expressing eukaryotic recombinants and the changes of telomerase activation in telomerase-positive cell line Hela. Methods： According to the sequence of NBS1 mRNA, the NBS1 pre-microRNA was designed and synthesized, then cloned into the GFP reporter pcDNA6.2-GW/EmGFP-miR vector and transfected into Hela cells. The integrity of the insert fragment was verified through colony PCR and sequencing analysis. The NBS1 gene expression of NBS1 microRNA recombinants was detected by Real-Time PCR and western blot. Telomerase activity in Hela cells was assayed by TRAP-PCR-EB. Results： Sequences of insert fragment in microRNA expressing recombinants were correct. The NBS1 gene expression was decreased, and the telomerase activation of Hela cell reduced. Conclusion： NBS1 microRNA inhibits NBS1 gene expression, and depresses telomerase activation of Hela cells. This confirms that there is relevance between NBS 1 gene and telomerase activity.
基金supported by grants from National Basic Research Program of China(2005CB522404,2006CB910506)The Program for Changjiang Scholars and Innovative Research Team(PCSIRT)in Universities(IRT0519)The National Natural Science Foundation of China(30771232,30671184)~~
基金supported by grants from the National Key Research and Development Program of China 2016YFA0500304 to T.K.the National Nature Science Foundation in China(NSFC)81772922 to Y.W.,81702890 to X.W.,and 81530081 and 31571395 to T.K.+2 种基金Guangdong Natural Science Foundation Team Project(2014A030312015 to T.K)the Sci-Tech Project Foundation of Guangzhou City(201607020038 to T.K.)the Natural Science Foundation of Guangdong Province(2016A030310218 to Y.W.).
文摘Human single-stranded DNA-binding protein 1(hSSB1)is required for the efficient recruitment of the MRN complex to DNA doublestrand breaks and is essential for the maintenance of genome integrity.However,the mechanism by which hSSB1 recruits NBS1 remains elusive.Here,we determined that hSSB1 undergoes SUMOylation at both K79 and K94 under normal conditions and that this modification is dramatically enhanced in response to DNA damage.SUMOylation of hSSB1,which is specifically fine-tuned by PIAS2α,and SENP2,not only stabilizes the protein but also enhances the recruitment of NBS1 to DNA damage sites.Cells with defective hSSB1 SUMOylation are sensitive to ionizing radiation,and global inhibition of SUMOylation by either knocking out UBC9 or adding SUMOylation inhibitors significantly enhances the sensitivity of cancer cells to etoposide.Our findings reveal that SUMOylation,as a novel posttranslational modification of hSSB1,is critical for the functions of this protein,indicating that the use of SUMOylation inhibitors(e.g.,2-D08 and ML-792)may be a new strategy that would benefit cancer patients being treated with chemo-or radiotherapy.
