Liang F, Jasin M
Molecular Biology Program, Sloan-Kettering Institute and Cornell University Graduate School of Medical Sciences, New York, New York 10021, USA.
J Biol Chem. 1996 Jun 14;271(24):14405-11. doi: 10.1074/jbc.271.24.14405.
Mammalian cells possess a protein complex, termed DNA-PK, which binds to DNA double strand breaks in vitro. The complex consists of the heterodimeric Ku autoantigen and a DNA-dependent protein kinase, DNA-PKcs. Cell lines that are deficient for components of this complex are sensitive to ionizing radiation and have impaired V(D)J recombination, a site-specific recombination process. We have tested these cell lines for their ability to repair double strand breaks in transfected DNA. The xrs-6 cell line, which is deficient for the 80-kDa subunit of the Ku autoantigen, exhibited reduced stability of transfected DNA. Prior to obvious reductions in DNA stability, the levels of homologous recombination and DNA end joining were unaffected. However, the recovery of end joining products with precisely joined ends was reduced, with a concomitant increase in products containing deletions. Unlike the Ku80-deficient cells, no reduction in DNA stability was detected in DNA-PKcs-deficient scid cells. Scid cells also exhibited normal levels of homologous recombination and DNA end joining. These experiments implicate the Ku autoantigen, but not DNA-PKcs, in a direct role in protecting DNA ends from degradation.
哺乳动物细胞拥有一种称为DNA-PK的蛋白质复合物，它在体外能与DNA双链断裂处结合。该复合物由异二聚体Ku自身抗原和一种依赖DNA的蛋白激酶DNA-PKcs组成。缺乏这种复合物成分的细胞系对电离辐射敏感，且V(D)J重组（一种位点特异性重组过程）受损。我们检测了这些细胞系修复转染DNA中双链断裂的能力。xrs-6细胞系缺乏Ku自身抗原的80 kDa亚基，其转染DNA的稳定性降低。在DNA稳定性明显降低之前，同源重组和DNA末端连接水平未受影响。然而，精确连接末端的末端连接产物的恢复减少，同时含有缺失的产物增加。与缺乏Ku80的细胞不同，在缺乏DNA-PKcs的scid细胞中未检测到DNA稳定性降低。scid细胞的同源重组和DNA末端连接水平也正常。这些实验表明，Ku自身抗原而非DNA-PKcs在直接保护DNA末端免受降解方面发挥作用。
