| Summary: | 碩士 === 國立清華大學 === 生命科學系 === 92 === In mammalian cells, there are two different pathways for repairing DNA double strand breaks (DSBs). One is homologous recombination, and the other is non-homologous end joining. In mammalian cells, DSBs repair proceeds predominantly by non-homologous end joining. In non-homologous end joining pathway, DNA-dependent protein kinase (DNA-PK) takes on an important responsibility. Ku protein is a subunit in DNA-PK. Ku protein is a heterodimer composed of Ku70 and Ku80. Ku protein can interact with double-stranded DNA ends. Ku protein binds DNA ends and recruits the catalytic subunit of DNA-PK (DNA-PKcs) to DNA in order to assemble an active complex. DNA-PK active complex recruits DNA ligase Ⅳ and XRCC4 to repair DSBs. In mammalian cells, deficiency of either Ku protein or DNA-PKcs is the major reason to cause DNA double strand break repair failure.
In this study, we demonstrated that Ku protein could not only bind DNA ends but also can transfer between two DNA molecules. The transfer of Ku is not to release from one DNA and bind to another DNA. Instead, Ku protein can transfer directly between two DNA molecules with homologous cohesive ends. Besides, increasing concentration of sodium chloride in the reaction, Ku protein can transfer between DNA with non-homologous ends. However, Ku protein can not transfer from DNA with random sequence (general DNA molecule) to a poly-dAdT fragment even with homologous cohesive ends. But, Ku protein can transfer from a poly-dAdT fragment to a general DNA molecule with homologous cohesive ends. Interestingly, increasing random DNA sequence in the poly-dAdT fragment, we observed Ku protein can transfer to the poly-dAdT fragment. In conclusions, we suggest from these results that there are two steps in the direct transfer of Ku protein. The first step is the formation of a DNA-Ku-DNA complex. In this step, it is required that the two DNA molecules have homologous cohesive ends under low salt condition but not under high salt condition. In the second step, Ku protein moves from one DNA molecule to another one. In this step, the dinection of movement of Ku under certain situation is influenced by DNA sequences.
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