CRISPR screens unveil nutrient-dependent lysosomal and mitochondrial nodes impacting intestinal tissue-resident memory CD8+ T cell formation.

Nutrient availability and organelle biology direct tissue homeostasis and cell fate, but how these processes orchestrate tissue immunity remains poorly defined. Here, using in vivo CRISPR-Cas9 screens, we uncovered organelle signaling and metabolic processes shaping CD8+ tissue-resident memory T (T RM) cell development. T RM cells depended on mitochondrial translation and respiration. Conversely, three nutrient-dependent lysosomal signaling nodes—Flcn, Ragulator, and Rag GTPases—inhibited intestinal T RM cell formation. Depleting these molecules or amino acids activated the transcription factor Tfeb, thereby linking nutrient stress to T RM programming. Further, Flcn deficiency promoted protective T RM cell responses in the small intestine. Mechanistically, the Flcn-Tfeb axis restrained retinoic acid-induced CCR9 expression for migration and transforming growth factor β (TGF-β)-mediated programming for lineage differentiation. Genetic interaction screening revealed that the mitochondrial protein Mrpl52 enabled early T RM cell formation, while Acss1 controlled T RM cell development under Flcn deficiency-associated lysosomal dysregulation. Thus, the interplay between nutrients, organelle signaling, and metabolic adaptation dictates tissue immunity. [Display omitted] • Systematic discovery of mitochondrial and lysosomal pathways in CD8+ T RM formation • Lysosomal signaling and amino acids shape Tfeb-driven T RM development • Small intestine-specific T RM programming is impeded by the Flcn-Tfeb signaling axis • Acss1 and Mrpl52 empower early siIEL T RM formation in distinct contexts How organelle signaling and metabolic adaptation orchestrate tissue-resident CD8+ T (T RM) cell development remains poorly defined. Here, Raynor et al. establish three nutrient-dependent lysosomal signaling nodes as negative regulators of T RM differentiation in the small intestine. These results uncover mechanisms dictating T RM cell quality and quantity for tissue immunity. [ABSTRACT FROM AUTHOR]