Fig. 4: Stimulation of MutSγ-MutLγ by nuclease-proficient EXO1 variants.

a A schematic overview of catalytically active EXO1 full-length and truncation variants used in this study. The N-terminal catalytic domain is shown in purple, and the C-terminal tail is shown in magenta. b Recombinant catalytically active full-length EXO1 and truncation mutants used in this study. The polyacrylamide gel was stained with Coomassie brilliant blue. A representative of two independent experiments. c Nuclease assays using 3’-labeled 50 bp dsDNA with indicated EXO1 variants. Top, quantification. Averages shown, n = 5 independent experiments; error bars, SEM. Bottom, a representative gel. The red asterisk indicates the position of the radioactive label. d Stimulation of MutLγ-MutSγ complex nicking activity by wild-type EXO1 or nuclease-dead EXO1 (D173A). Reactions were carried out in the presence of 0.6 mM Mn²⁺ and in the absence of RFC-PCNA (left) or in the presence of 0.6 mM Mg²⁺ and RFC-PCNA (right). Nuclease-dead MutLγ (MutLγ-3ND, [D1223N, Q1224K, E1229K]) was used as a control (lanes 4, 7, 11 and 14). Top, quantification; averages shown, n = 4 for experiments in Mn²⁺ and n = 7 for experiments in Mg²⁺; error bars, SEM. Bottom, representative gels. e Stimulation of MutLγ-MutSγ complex nicking activity by nuclease-proficient full-length EXO1, EXO1 1-390 and EXO 1-352. Reactions were carried out in the presence of 0.6 mM Mn²⁺ in the absence of RFC-PCNA (left) or in the presence of 0.6 mM Mg²⁺ and RFC-PCNA (right). Nuclease-dead MutLγ (MutLγ-3ND, [D1223N, Q1224K, E1229K]) was used as a control (lanes 6 and 12). Top, quantification; averages shown, n = 4 independent experiments; error bars, SEM. Bottom, representative gels, two-tailed t-test. Source data are provided as a Source Data file.