Expression and activity of DNA-dependent protein kinase in normal human leukocytes.
Expression and activity of DNA-dependent protein kinase in normal human leukocytes.
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发表时间：
2000-09
影响因子：
2
通讯作者：
A. Eriksson;R. Lewensohn;A. Nilsson
中科院分区：
文献类型：
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作者：
A. Eriksson;R. Lewensohn;A. Nilsson
背景 DNA-PK 是一种丝氨酸/苏氨酸蛋白激酶，已被证明在 V(D)J 重组、Ig 转换和 DNA 双链断裂 (DSB) 修复中发挥重要作用。我们研究了 B 和 T 淋巴细胞、单核细胞和粒细胞的全细胞提取物中的 DNA-PK，以及佛波肉豆蔻酸乙酸酯 (PMA) 和刀豆球蛋白 A (Con A) 对 B 和 T 淋巴细胞中 DNA-PK 的影响。材料和方法 使用 E-rosette 技术从人外周血中分离淋巴细胞，并使用蛋白质印迹、DNA-PK 激酶活性测定和流式细胞术分析 DNA-PK。结果 DNA-PK 活性水平和含量在白细胞之间存在差异。当细胞进入 S 期时，PMA 和 Con A 刺激持续增加 B 淋巴细胞中的 DNA-PK 活性和含量。尽管细胞在 PMA 和 Con A 刺激后进入 S 期，但 T 淋巴细胞中通常看不到上调。结论 有丝分裂刺激上调 B 淋巴细胞中 DNA-PK 活性和蛋白质表达。 T 淋巴细胞的 DNA-PK 显示个体间差异。 DNA-PK 的调节并不严格依赖于所有细胞类型的细胞周期。
BACKGROUND DNA-PK is a serine/threonine protein kinase that has been shown to play an important role in V(D)J recombination, Ig switching and DNA double strand break (DSB) repair. We have investigated DNA-PK in whole cell extracts from B and T lymphocytes, monocytes and granulocytes and the effects of phorbolmyristate acetate (PMA) and Concanavalin A (Con A) on DNA-PK in B and T lymphocytes. MATERIALS AND METHODS Lymphocytes were separated from human peripheral blood using the E-rosette technique and DNA-PK was analysed using Western blot, DNA-PK kinase activity assay and flow cytometry. RESULTS The levels of DNA-PK activity and content varied among the leukocytes. PMA and Con A stimulation consistently increased both DNA-PK activity and content in B lymphocytes as cells entered the S phase. Up-regulation could normally not be seen in T lymphocytes, although cells entered the S phase following PMA and Con A stimulation. CONCLUSION Mitogenic stimulation up-regulates DNA-PK activity and protein expression in B lymphocytes. T lymphocytes show interindividual differences in DNA-PK. Regulation of DNA-PK is not strictly cell cycle dependent for all cell types.