Methyl-HLH-30/TFEB orchestrates holistic intestinal recuperation against bacterial pore-forming toxin intoxication via TTC38 in metazoans.

Effector-triggered immunity (ETI) safeguards intestinal epithelial cells (IECs) by surveilling bacterial virulence effectors or the damage they cause, such as pore-forming toxins (PFTs) that breach plasma membrane integrity (PMI). However, how metazoan ETI regulators guard IEC homeostasis compromised by PFT in vivo remains elusive. Moreover, while the molecular architectures of ETI are well-defined in plants, functionally analogous systems in animals are only beginning to emerge. Here, we unveil a conserved ETI guard system - the intestinal ETI regulator methyl-HLH-30/TFEB upregulates a novel intrinsic cellular defense (INCED) orchestrator Y54G11A.4/TTC38 - in Caenorhabditis elegans and human IECs. This conserved ETI system coordinates the restoration of PMI, microvilli architecture, and nutrient absorption concurrently by promoting phase condensation of the PTRN-1/CAMSAP-dependent guard program through an HSPA/HSP70-HSP90 chaperone system. Together, our findings reveal an intestinal ETI guard system that couples PFT-damage sensing to comprehensive epithelial recuperation, underscoring its physiological relevance for mucosal homeostasis across metazoans.Abbreviations and acronyms B. thuringiensis (Bt): Bacillus thuringiensis; CAMSAP: calmodulin regulated spectrin associated protein; C. elegans: Caenorhabditis elegans; Cry toxins: crystal toxins; ETI: effector-triggered immunity; FRAP: fluorescence recovery after photobleaching; FRET: Förster/fluorescence resonance energy transfer; GFP: green fluorescent protein; hlh/HLH: helix loop helix; HSP: heat shock protein; IDRs: intrinsically disordered regions; IECs: intestinal epithelial cells; INCED: intrinsic cellular defense; IPTG: isopropyl-β-D-1-thiogalactopyranoside; lgg/LGG: LC3 and GABARAP family; ncMTOC: non-centrosomal microtubule-organizing center; LLPS: liquid-liquid phase separation; PFTs: pore-forming toxins; PI: propidium iodide; PMI: plasma membrane integrity; PTMs: post-translational modifications; qRT-PCR: quantitative real-time PCR; RNAi: ribonucleic acid interference; SEM: scanning electron microscopy; SLO: streptolysin O; SQST-1/SQSTM1/p62: sequestosome related 1; TFEB: transcription factor EB; TPR: tetratricopeptide repeat motifs; TTC38: tetratricopeptide repeat domain 38.