Abstract Mitochondria are dynamic organelles that must continually adapt and respond to cellular stress. Recent studies demonstrated that mitochondrial stress can be relayed from mitochondria to the cytosol by the release of a C-terminal proteolytic fragment of DELE1 that binds to the eIF2α kinase HRI to initiate integrate stress response (ISR) signaling. Here, we report the cryo-electron microscopy structure of the active, C-terminal cleavage product of human DELE1 at ∼ 3.8 A° resolution. Our structure reveals that DELE1 assembles into a high-order oligomer that is observed both in vitro and in mammalian cells. Structurally, the oligomer consists of eight DELE1 monomers that assemble with D4 symmetry via two sets of distinct hydrophobic inter-subunit interactions. We identified the key residues involved in DELE1 oligomerization, and confirmed their role in stabilizing the octamer in vitro and in cells using mutagenesis. Further, we show that assembly impaired DELE1 mutants are compromised in their ability to induce ISR activation in cell culture models. Together, our findings provide molecular insights into the activity of DELE1 and how it signals to promote ISR activity following mitochondrial insult.

DELE1 oligomerization promotes integrated stress response activation