国际放射医学核医学杂志
國際放射醫學覈醫學雜誌
국제방사의학핵의학잡지
INTERNATIONAL JOURNAL OF RADIATION MEDICINE AND NUCLEAR MEDICINE
2012年
1期
30-34
,共5页
杨天一%张士猛%秦夏%李兵%刘晓丹%周平坤
楊天一%張士猛%秦夏%李兵%劉曉丹%週平坤
양천일%장사맹%진하%리병%류효단%주평곤
反式激活%辐射效应%DNA依赖蛋白激酶催化亚单位基因启动子%染色体重构
反式激活%輻射效應%DNA依賴蛋白激酶催化亞單位基因啟動子%染色體重構
반식격활%복사효응%DNA의뢰단백격매최화아단위기인계동자%염색체중구
Trans-activation%Radiation effects%DNA dependent protein kiuase catalytic subunit promoter%Chromatin remodeling
目的 检测人类免疫缺陷病毒转录反式激活因子( TAT)对DNA依赖蛋白激酶催化亚单位( DN A-PKcs)基因启动子活性的影响.方法 用PCR方法克隆DNA-PKcs基因启动子的系列截短体,通过DNA重组构建pGL3-basic-DN A-PKcs肩动子报告质粒载体,通过双荧光素酶报告基因检测DNA-PKcs基因的肩动子活性,采用基于乳糖抑制子和乳糖操纵子并结合绿色荧光蛋白分子荧光的大规模染色质松弛报告系统观察染色质的重构活性,并研究了电离辐射对TAT参与染色体重构的影响.结果构建了DNA-PKcs启动子(全长区域为-939 bp～-1 bp)的系列截短体的报告质粒载体,鉴定出其核心区域为-64 bp～-1 bp.TAT能够抑制DNA-PKcs基因启动子的转录活性.TAT具有大规模的染色体松弛活性,电离辐射能抑制TAT参与染色体重构的作用.结论 TAT能抑制DNA修复基因DNA-PKcs的启动子活性,电离辐射能抑制TAT参与染色体重构的作用.
目的 檢測人類免疫缺陷病毒轉錄反式激活因子( TAT)對DNA依賴蛋白激酶催化亞單位( DN A-PKcs)基因啟動子活性的影響.方法 用PCR方法剋隆DNA-PKcs基因啟動子的繫列截短體,通過DNA重組構建pGL3-basic-DN A-PKcs肩動子報告質粒載體,通過雙熒光素酶報告基因檢測DNA-PKcs基因的肩動子活性,採用基于乳糖抑製子和乳糖操縱子併結閤綠色熒光蛋白分子熒光的大規模染色質鬆弛報告繫統觀察染色質的重構活性,併研究瞭電離輻射對TAT參與染色體重構的影響.結果構建瞭DNA-PKcs啟動子(全長區域為-939 bp～-1 bp)的繫列截短體的報告質粒載體,鑒定齣其覈心區域為-64 bp～-1 bp.TAT能夠抑製DNA-PKcs基因啟動子的轉錄活性.TAT具有大規模的染色體鬆弛活性,電離輻射能抑製TAT參與染色體重構的作用.結論 TAT能抑製DNA脩複基因DNA-PKcs的啟動子活性,電離輻射能抑製TAT參與染色體重構的作用.
목적 검측인류면역결함병독전록반식격활인자( TAT)대DNA의뢰단백격매최화아단위( DN A-PKcs)기인계동자활성적영향.방법 용PCR방법극륭DNA-PKcs기인계동자적계렬절단체,통과DNA중조구건pGL3-basic-DN A-PKcs견동자보고질립재체,통과쌍형광소매보고기인검측DNA-PKcs기인적견동자활성,채용기우유당억제자화유당조종자병결합록색형광단백분자형광적대규모염색질송이보고계통관찰염색질적중구활성,병연구료전리복사대TAT삼여염색체중구적영향.결과구건료DNA-PKcs계동자(전장구역위-939 bp～-1 bp)적계렬절단체적보고질립재체,감정출기핵심구역위-64 bp～-1 bp.TAT능구억제DNA-PKcs기인계동자적전록활성.TAT구유대규모적염색체송이활성,전리복사능억제TAT삼여염색체중구적작용.결론 TAT능억제DNA수복기인DNA-PKcs적계동자활성,전리복사능억제TAT삼여염색체중구적작용.
Objective To explore the influence of human immunodeficiency virus transactivator of transcription(TAT) on the promoter activity of DNA dependent protein kinase catalytic subunit (DNA-PKcs).Methods The truncated promoters of DNA-PKcs were cloned by PCR from the template DNA from HeLa genomic DNA,and the pGL3-basic-DNA-PKcs promoter reporter plasmids were constructed.The activity of DNA-PKcs promoters was detected by dual-luciferase reporter assay system.A Lac-repressor and Lacoperator based green fluorescent protein imaging system was used to assay the chromatin remodeling activity.Results A series of reporter plasmids harboring the truncated promoters of DNA-PKcs from -939 bp to -1 bp were constructed.The sequence of -64 bp to-1 bp was identified as a critical element for the activity of DNA-PKes promoter.TAT can suppress the activity of DNA-PKcs promoter.TAT participates in the regulation of the large scale chromatin relaxation.Ionizing radiation attenuates the activity of TAT played in the chromatin remodeling.Conclusion TAT represses the promoter activity of DNA repair protein DNA-PKcs,and also play a role of large scale chromatin remodeling which can te attenuated by ionizing radiation.
