STYK1 interacts with the PtdIns3K-C1 complex. (A) HeLa cells were simultaneously transfected with four plasmids containing Flag-STYK1, HA-PIK3C3, HA-BECN1 and MYC-ATG14 for 48 h, then immunoprecipitated with an anti-Flag antibody and subjected to SDS-PAGE, followed by Coomassie blue staining. (B) The interaction between Flag-STYK1 and the components of the PtdIns3K-C1 complex in HeLa cells. The cell lysates were used for IP and western blotting with the indicated antibodies. (C) The interaction between endogenous BECN1 and Flag-STYK1. (D and E) The interactions between endogenous BECN1, ATG14 and endogenous STYK1. (F) The interaction between Flag-STYK1 and MYC-ATG14 in HEK293T cells. (G) Colocalization between STYK1-mCherry and BECN1, PIK3C3, PIK3R4. Scale bars: 10 µm. (H) Two-step co-IP assay in HEK293T cells to determine the complex containing STYK1, BECN1, and ATG14. The procedures of the two-step co-IP are outlined at the top. The samples were subjected to western blotting with the indicated antibodies.