biostudies-literatureChen MNINDS NIH HHSNIGMS NIH HHS1596-1605https://www.ebi.ac.uk/biostudies/studies/S-EPMC12440827biostudies-literatureUnknown32(9)The Unc-51-like kinase protein kinase complex (ULK1C) is the most upstream and central player in the initiation of macroautophagy in mammals. Here, we determined the cryo-electron microscopy structure of the human ULK1C core at amino-acid-level resolution. We also determined a moderate-resolution structure of the ULK1C core in complex with another autophagy core complex, the class III phosphatidylinositol 3-OH kinase complex I (PI3KC3-C1). We show that the two complexes coassemble through extensive contacts between the FIP200 scaffold subunit of ULK1C and the VPS15, ATG14 and BECN1 subunits of PI3KC3-C1. The FIP200:ATG13:ULK1 core of ULK1C undergoes a rearrangement from 2:1:1 to 2:2:2 stoichiometry in the presence of PI3KC3-C1. This suggests a structural mechanism for the initiation of autNature structural & molecular biologyStructure and activation of the human autophagy-initiating ULK1C:PI3KC3-C1 supercomplex.PMC12440827R35 GM136414R01 NS134598Ren XCook ASILazarou MHurley JHChen MNguyen TNZhao YKhuu GYildiz AfalseStructure and activation of the human autophagy-initiating ULK1C:PI3KC3-C1 supercomplex.The Unc-51-like kinase protein kinase complex (ULK1C) is the most upstream and central player in the initiation of macroautophagy in mammals. Here, we determined the cryo-electron microscopy structure of the human ULK1C core at amino-acid-level resolution. We also determined a moderate-resolution structure of the ULK1C core in complex with another autophagy core complex, the class III phosphatidylinositol 3-OH kinase complex I (PI3KC3-C1). We show that the two complexes coassemble through extensive contacts between the FIP200 scaffold subunit of ULK1C and the VPS15, ATG14 and BECN1 subunits of PI3KC3-C1. The FIP200:ATG13:ULK1 core of ULK1C undergoes a rearrangement from 2:1:1 to 2:2:2 stoichiometry in the presence of PI3KC3-C1. This suggests a structural mechanism for the initiation of aut2025-01-01T00:00:00Z2025 Sep2026-06-03T08:00:57.535Z2026-04-26T03:12:05.58ZS-EPMC124408274044231610.1038/s41594-025-01557-x