Thymine DNA glycosylase combines sliding, hopping, and nucleosome interactions to efficiently search for 5-formylcytosine
Brittani L Schnable, Matthew A. Schaich, Vera Roginskaya, Liam P. Leary, Tyler Weaver, Bret Freudenthal, Alexander C. Drohat, Bennett Van Houten
Abstract
Base excision repair is the main pathway involved in active DNA demethylation. 5-formylcytosine and 5-carboxylcytosine, two oxidized moieties of methylated cytosine, are recognized and removed by thymine DNA glycosylase (TDG) to generate an abasic site. Using single molecule fluorescence experiments, we study TDG in the presence and absence of 5-formylcytosine. TDG exhibits multiple modes of linear diffusion, including hopping and sliding, in search of base modifications. TDG active site variants and truncated N-terminus, reveals these variants alter base modification search and recognition mechanism of TDG. On DNA containing an undamaged nucleosome, TDG is found to either bypass, colocalize with, or encounter but not bypass the nucleosome. Truncating the N-terminus reduces the number of interactions with the nucleosome. Our findings provide mechanistic insights into how TDG searches for modified DNA bases in chromatin. Thymine DNA glycosylase is an essential DNA repair enzyme involved in oxidative demethylation of 5- methylC. In this study the authors have tracked specific and non-specific DNA binding of TDG at the single molecule level. TDG is found to hop and slide on DNA and found to interact with nucleosomes.