FIGURE 2.

Roles of DNA Pol β in in vitro LTR DNA gap repair activity of nuclear extracts from human primary macrophages and activated CD4⁺ T cells. A, nuclear extracts from human macrophages and activated CD4⁺ T cells from two donors (d1 and d2) were normalized to 4 μg/μl total protein and immunodepleted (Im. Depl.) for Pol β or control anti-α-tubulin antibody. The depleted extracts were incubated with substrate i for 30 min with 250 μm dNTPs (upper panel). The depleted extract was Western-blotted (WB) for the presence of either DNA Pol β or actin (lower panels). The lanes in the lower panels correspond to the immunodepletion indicated in the upper panel. The 24-mer gap-filled product (B) and the fully repaired 50-mer product (C) in the reactions with macrophages and T cells immunodepleted with DNA Pol β or anti-α-tubulin (Tub) antibody were quantitated against the total product. As shown in Fig. 2 (B and C, see single and double asterisks), there was a significant reduction in both the 24-mer gap-filled product and the 50-mer ligated product.