PINK1 physically interacts with IRAK1 in HEK293 cells. (A) HEK293 cells were transfected for 48 hours with Myc-PINK1 and/or Flag-IRAK1. Cell lysates were immunoprecipitated with the c-Myc antibody, followed by immunoblotting with the Flag antibody. Expression of the transiently transfected proteins was identified by immunoblotting with the indicated antibodies. *IgG heavy chains. Actin was used as a loading control. (B) HEK293 cell lysates were immunoprecipitated with the IRAK1 antibody, followed by immunoblotting with the PINK1 antibody. As a control, cell lysates were immunoprecipitated with preimmune IgG. (C) HEK293 cells were fixed, permeabilized, and incubated for 24 hours with PINK1 or IRAK1 antibodies. Cells were stained with TRITC-conjugated or FITC-conjugated secondary antibodies and 4′,6′-diamidino-2-phenylindole (DAPI). Immunostained cells were examined using confocal microscopy. Scale bar = 20 μm. IRAK1, IL-1 receptor-associated kinase 1; PINK1,
PTEN-induced putative kinase 1.