In order to gain an in depth molecular understanding Moreover, exact mechanistic role of histone PTM on BER inside cell is unclear. In order to examine the influence of histone acetylation on the initial steps of BER, we assembled nucleosome arrays consisting of homogeneously acetylated histone H3 (H3K18 and H3K27) and measured the repair of a site-specifically positioned 2′-deoxyuridine (dU). We find that H3K18ac and H3K27ac differentially influence the combined activities of UDG/APE1 on chromatin in a context-dependent manner, suggesting that acetylated lysine residues on the H3 tail domain play distinct roles in regulating the initial steps of BER.2 Overall, I will highlights the value of performing detailed biochemical studies on precisely modified chromatin substrates for investigating DNA damage repair in chromatin biology and provides our finding on how higher-order chromatin structure and histone modification regulates BER on chromatin. of how chromatin structure and histone PTM regulates the access, detailed in vitro biochemical and biophysical studies are required. However, because of challenges associated with reconstituting nucleosome arrays containing site-specifically positioned modifications, such studies have been limited to the use of mono- and dinucleosomes as model in vitro substrates, which are incapable of folding into native chromatin structures. To address this issue, we developed a straightforward and general approach for assembling chemically defined oligonucleosome arrays (i.e., designer chromatin) containing site-specifically modified DNA and histone PTM. Using this approach, we prepared several oligonucleosome substrates containing precisely positioned 2′-deoxyuridine (dU) residues and examined the efficiency of base excision repair (BER) within several distinct chromatin architectures. We show that, depending on the translational position of the lesion, BER efficacy can be either inhibited by as much as 20-fold or accelerated by more than 5-fold within compact chromatin (i.e., the 30 nm fiber) relative to naked DNA. Moreover, we demonstrate the first direct evidence that internucleosome interactions play an important role in regulating BER within higher-order chromatin structures.