Figure 5: PAXX promotes the assembly of XLF into the DNA end-bound Ku complex. | Nature Communications

Figure 5: PAXX promotes the assembly of XLF into the DNA end-bound Ku complex.

From: Interactome analysis identifies a new paralogue of XRCC4 in non-homologous end joining DNA repair pathway

Figure 5

(a) Immunoblotting showing Flag-PAXX with or without EtBr. (b) Electrophoretic mobility shift assay (EMSA) showing that PAXX does not bind DNA. Reactions contain 5 nM Cy3-labelled 30-bp dsDNA and 10, 30, 100 or 300 nM PAXX. (c) EMSA assay showing that PAXX but not its mutant F201A supershifts the DNA–Ku complex. Reactions were performed as in b, excluding the presence of Ku in the indicated wells. The composition of the protein–DNA complexes is indicated on the right side. (d) EMSA showing that PAXX and XLF mutually promote their recruitment to the DNA–Ku complex. Reactions contain 5 nM Cy3-labelled 30-bp dsDNA and the indicated proteins. The series of concentrations was 50, 100, 200 and 400 nM for XLF, and 30, 100 and 300 nM for PAXX. (e,f) Quantifications of reduced complexes. The reduced complexes in d, lanes 6–10 (f) and lanes 11–15 (e) were quantified. The complex quantities in lanes 6 and 11 were set at 100% for the respective experiments.

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