Structure and activation of the human autophagy-initiating ULK1C:PI3KC3-C1 supercomplex

Abstract

The unc-51-like kinase protein kinase complex (ULK1C) and the class III phosphatidylinositol (PI) 3-kinase complex I (PI3KC3-C1) are the most upstream and central players in the initiation of macroautophagy. We found a direct physical interaction between the two complexes. The cryo-EM structures of the human ULK1C core and PI3KC3-C1 were determined at amino acid residue-level resolution, and used to interpret a moderate resolution structure of the ULK1C:PI3KC3-C1 supercomplex. The two complexes coassemble through extensive contacts between the FIP200 scaffold subunit of ULK1C and the VPS15 pseudokinase subunit of PI3KC3-C1. The presence of PI3KC3-C1 induces a rearrangement of ULK1C from a FIP200:ATG13:ULK1 2:1:1 to a 2:2:2 stoichiometry by dislocating an ATG13 loop (ATG13MR) from an inhibitory site on the dimeric FIP200 scaffold. This suggests a mechanism for the initiation of autophagy through PI3KC3-C1-induced dimerization of ULK1 as bound to FIP200, followed by an activating trans-autophosphorylation of ULK1.

Structural view on autophagosome formation

“…96 A, revealing the overall structure of the human PI-3 kinase complex (Fig. 4D) [106,118,119]. These studies unveiled the intricate architecture of this large complex (Fig.…”

“…Additionally, negative stain EM and biochemical data indicated that ULK1, ATG13, and FIP200 form a 1 : 1 : 2 complex, with the detailed structure revealed later using cryo-EM assisted with AlphaFold2 (Fig. 3J) [106].…”

“…In this super complex, PI3KC3-C1 induces a rearrangement of ULK1 by dislocating an ATG13 loop from an inhibitory site on the FIP200 dimer, changing the stoichiometry of ULK1, ATG13, and FIP200 in the complex from 1 : 1 : 2 to 2 : 2 : 2. This stoichiometric change was proposed to promote self-phosphorylation of ULK1 to activate its kinase activity [106]. Further investigation is needed to fully understand the biological significance of the formation of this super complex.…”

Epstein–Barr Virus Encoded BCL2, BHRF1, Downregulates Autophagy by Noncanonical Binding of BECN1

“…BECN1 is a key structural component of the class III phosphatidylinositol 3-kinase (PI3Kc3) complexes that are critical for autophagosome nucleation. , BECN1 is a conformationally flexible protein conserved in all eukaryotes that contains at least four domains: an N-terminal intrinsically disordered region (IDR), , a flexible helical domain (FHD), , a coiled-coil domain (CCD), , and a β–α repeated, autophagy-specific domain (BARAD). , The residues comprising these domains in human BECN1 are IDR, residues 1–140, FHD, residues 141–171, CCD, residues 175–265, and BARAD, residues 265–450 . The BARAD bears an aromatic finger comprising residues F359, F360, and W361 that enables BECN1 to interact with membranes. , The BECN1 IDR includes a BH3D, comprising residues 105–130, classifying it as a BH3-only protein .…”