Interaction between hMYH and hRad9. (A) HEK293 cells were transfected with c-myc and FLAG-tagged hRad9, or c-myc-tagged hMYH and FLAG-tagged-hRad9 as indicated. Co-IP was performed with anti-c-myc antibody, and the immunoprecipitates were immunoblotted with anti-FLAG and anti-c-myc antibodies. (B) Lysates from untransfected HEK293 cells were used for the immunoprecipitation of hMYH with an anti-hMYH antibody. Immunoblotting was performed using anti-hMYH and anti-hRad9 antibodies. Lane 1: input sample, lanes 2 and 3: proteins immunoprecipitated with mouse IgG or anti-hMYH antibody. (C) HEK293 cells were transfected with siHus1 or siGFP. Total cell lysates were used for co-IP with hRad9 antibody. (D) Pull-down of purified His-hRad9 with GST-hMYH bound to glutathione-Sepharose beads. Lane 1: purified His-hRad9, lanes 2 and 3: pull-downs with GST and GST-hMYH, respectively.