| 總結: | <p>Living organisms have evolved multiple repair pathways that recognize and tackle different types of DNA damage. One such pathway is the Fanconi anemia DNA repair pathway which is able to repair DNA interstrand crosslinks (ICLs) – deleterious lesions arising by a covalent binding of the opposite DNA strands through a linker molecule. Of central importance to the ICL repair lies the FANCD2/FANCI heterodimer which is recruited to the site of the lesion and is subsequently monoubiquitinated by the Fanconi anemia core complex. These two steps are essential for the pathway to proceed and if they fail, so fails the entire repair. Due to its importance, FANCD2/FANCI is highly regulated by numerous post-translational modifications, phosphorylation being the best characterized.</p>
<p>In the work presented herein, I focus on FANCD2 phosphorylation and its linkage to ATR. In the first two chapters I have demonstrated that FANCD2 is activated by phosphorylation at multiple residues which enables its recruitment to ICLs as well as its monoubiquitination. This phosphorylation occurs in two steps. First, before its recruitment to ICLs, and subsequently between the recruitment and the monoubiquitination step. These data also suggest that FANCD2 recruitment and monoubiquitination are partially facilitated by ATR. In the third chapter I used a Single Particle Tracking technique to start characterising the kinetics of FANCD2 recruitment. It was shown that an increased number of FANCD2 molecules binds to chromatin after the introduction of ICLs. An increased number of bound FANCD2 was observed also in the unperturbed S phase cells, and depletion of FANCI nullifies this increase. Finally, the mean retention time of FANCD2 was determined to be approximately 30 seconds in unperturbed cells.</p>
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