TECPR1 conjugates LC3 to damaged endomembranes upon detection of sphingomyelin exposure

Abstract

Invasive bacteria enter the cytosol of host cells through initial uptake into bacteria-containing vacuoles (BCVs) and subsequent rupture of the BCV membrane, thereby exposing to the cytosol intraluminal, otherwise shielded danger signals such as glycans and sphingomyelin. The detection of glycans by galectin-8 triggers anti-bacterial autophagy, but how cells sense and respond to cytosolically exposed sphingomyelin remains unknown. Here, we identify TECPR1 (tectonin beta-propeller repeat containing 1) as a receptor for cytosolically exposed sphingomyelin, which recruits ATG5 into an E3 ligase complex that mediates lipid conjugation of LC3 independently of ATG16L1. TECPR1 binds sphingomyelin through its N-terminal DysF domain (N'DysF), a feature not shared by other mammalian DysF domains. Solving the crystal structure of N'DysF, we identified key residues required for the interaction, including a solvent-exposed tryptophan (W154) essential for binding to sphingomyelin-positive membranes and the conjugation of LC3 to lipids. Specificity of the ATG5/ATG12-E3 ligase responsible for the conjugation of LC3 is therefore conferred by interchangeable receptor subunits, that is, the canonical ATG16L1 and the sphingomyelin-specific TECPR1, in an arrangement reminiscent of certain multi-subunit ubiquitin E3 ligases.

ESCRT recruitment to damaged lysosomes is dependent on the ATG8 E3-like ligases

“…Ca 2+ leakage will induce the scrambling of sphingomyelin (SM) from the luminal to cytoplasmic membrane surface of the lysosomal membrane (24). SM will subsequently recruit the TECPR1-ATG5-ATG12 E3-like ligase to the damaged membrane (21)(22)(23). Here we show that ATG8 E3-like ligase translocation is an essential prerequisite to ESCRT recruitment, thereby placing the ATG8 E3-like ligases in the role of damage sensors for the ESCRT-mediated membrane repair pathway.…”

“…We and others recently identified a TECPR1-dependent autophagy E3-like ligase complex which functions independently of ATG16L1 to regulate unconventional ATG8 lipidation at damaged lysosomal membranes (21)(22)(23). Double knockout of both ATG16L1 and TECPR1 was shown to compromise the repair of damaged membranes, suggesting a functionally redundant role for both E3-like ligase complexes in the repair process (21).…”

An ATG12‐ATG5‐TECPR1 E3‐like complex regulates unconventional LC3 lipidation at damaged lysosomes

“…Thus, we can conclude that the Nterminal dysferlin domain of TECPR1 is essential for its lysosomal recruitment in response to damage. In a companion manuscript, Boyle et al (2023) observed TECPR1 recruitment to damaged phagosomal membranes in response to bacterial infection. This recruitment is mediated by a sphingomyelin binding domain identified in the N-terminal dysferlin domain of TECPR1.…”

“…Lysosomal damage was recently shown to induce a rapid sphingomyelin translocation from the luminal to cytoplasmic membrane surface (Niekamp et al, 2022), suggesting sphingomyelin enrichment at damaged membranes could represent a conserved TECPR1 recruitment mechanism. To confirm, we introduced a point mutation at W154 of TECPR1, shown by Boyle et al (2023) to be essential for sphingomyelin binding. This single point mutation was sufficient to block TECPR1 lysosomal enrichment in response to LLOMe treatment (Fig EV1A -C).…”

A mechanism that transduces lysosomal damage signals to stress granule formation for cell survival

“…Chen et al, 2024; Radulovic et al, 2018; Skowyra et al, 2018). Severely damaged lysosomes can be removed by selective autophagy(Chauhan et al, 2016; Maejima et al, 2013), noncanonical autophagy(Boyle et al, 2023; Kaur et al, 2023), or lysosomal exocytosis(Wang et al, 2023). Master regulators mTORC1 (mechanistic target of rapamycin complex 1) and AMPK (AMP-activated protein kinase), located on lysosomes(Sancak et al, 2010; C.-S. Zhang et al, 2014), are finely tuned to respond to lysosomal damage, subsequently activating downstream processes e.g., autophagy and lysosomal biogenesis(Jia et al, 2018; Jia et al, 2020a, b; Jia et al, 2020c).…”