id: MODULE:phagolysosomal_cargo_degradation title: Phagolysosomal cargo degradation description: Once a phagosome has matured into a phagolysosome, acid hydrolases digest the engulfed particle and transporters in the phagolysosomal membrane return the products to the cytosol. DNase II finishes digesting the DNA of engulfed cells; cathepsins digest proteins; in amoebae that eat bacteria, lysozymes break down the bacterial cell wall. Amino acids, nucleosides, heme and cholesterol are then exported by dedicated transporters. Digestion is how phagotrophic protists feed and how animal phagocytes dispose of dead cells and microbes. Loss of a hydrolase or an exporter leaves undigested cargo, and undigested DNA in particular can trigger innate immune signalling. status: DRAFT scope: ABSTRACT references: - id: PMID:10716942 title: NUC-1, a caenorhabditis elegans DNase II homolog, functions in an intermediate step of DNA degradation during apoptosis. reference_review: relevance: HIGH correctness: VERIFIED review_notes: PubMed-verified; checked against the cached record. - id: PMID:12524536 title: Impaired thymic development in mouse embryos deficient in apoptotic DNA degradation. reference_review: relevance: HIGH correctness: VERIFIED review_notes: PubMed-verified; checked against the cached record. - id: PMID:11375492 title: Requirement of DNase II for definitive erythropoiesis in the mouse fetal liver. reference_review: relevance: HIGH correctness: VERIFIED review_notes: PubMed-verified; checked against the cached record. - id: PMID:12381665 title: Activation of the innate immunity in Drosophila by endogenous chromosomal DNA that escaped apoptotic degradation. reference_review: relevance: HIGH correctness: VERIFIED review_notes: PubMed-verified; checked against the cached record. - id: PMID:41826292 title: Microglial cathepsin B is necessary for neuronal efferocytosis in zebrafish and mice during brain development. reference_review: relevance: HIGH correctness: VERIFIED review_notes: PubMed-verified; checked against the cached record. - id: PMID:16424157 title: 'Cutting edge: a novel nonoxidative phagosomal mechanism exerted by cathepsin-D controls Listeria monocytogenes intracellular growth.' reference_review: relevance: HIGH correctness: VERIFIED review_notes: PubMed-verified; checked against the cached record. - id: PMID:22822152 title: LAAT-1 is the lysosomal lysine/arginine transporter that maintains amino acid homeostasis. reference_review: relevance: HIGH correctness: VERIFIED review_notes: PubMed-verified; checked against the cached record. - id: PMID:22174130 title: Equilibrative nucleoside transporter 3 deficiency perturbs lysosome function and macrophage homeostasis. reference_review: relevance: HIGH correctness: VERIFIED review_notes: PubMed-verified; checked against the cached record. - id: PMID:39547438 title: Deficiency of myeloid NPC1 exacerbates liver injury and fibrosis by impairing macrophage efferocytosis. reference_review: relevance: HIGH correctness: VERIFIED review_notes: PubMed-verified; checked against the cached record. - id: PMID:15640146 title: A Dictyostelium mutant with reduced lysozyme levels compensates by increased phagocytic activity. reference_review: relevance: HIGH correctness: VERIFIED review_notes: PubMed-verified; checked against the cached record. - id: PMID:23169667 title: Heptahelical protein PQLC2 is a lysosomal cationic amino acid exporter underlying the action of cysteamine in cystinosis therapy. reference_review: relevance: HIGH correctness: VERIFIED review_notes: PubMed-verified; checked against the cached record. notes: 'Boundary: starts with a degradative phagolysosome (the output of MODULE:phagosome_maturation) and ends with digestion products in the cytosol. Phagosome maturation, acidification and lysosome reformation are excluded. Signalling programmes triggered by digestion products are separate modules (MODULE:efferocytosis_induced_macrophage_proliferation, MODULE:efferocytosis_induced_anti_inflammatory_programme, MODULE:heme_induced_iron_recycling_macrophage_differentiation). The tryptophan exporter SLC36A4 is modelled in the anti-inflammatory module. Most of these hydrolases and transporters are general lysosomal machinery. They are included where there is loss-of-function evidence that engulfed cargo is not digested or its products accumulate. LIPA is left out because its efferocytosis evidence is pharmacological and its macrophage knockout is masked by cross-correction from other cells. Taxon scope: digestion of phagocytosed material is eukaryote-wide. DNase II activity is inferred at a eukaryote-level PAINT node. The evidence here comes from mouse, zebrafish, C. elegans, Drosophila and Dictyostelium.' module: id: phagolysosomal_cargo_degradation label: Phagolysosomal cargo degradation module_type: BIOLOGICAL_PROCESS concepts: - preferred_term: digestion of phagocytosed material in the phagolysosome notes: GO has no process term for this stage. GO:1900756 protein processing in phagocytic vesicle covers only the protein part. context: taxa: - preferred_term: Eukaryota term: id: NCBITaxon:2759 label: Eukaryota cell_types: - preferred_term: phagocyte term: id: CL:0000234 label: phagocyte cellular_components: - preferred_term: phagolysosome term: id: GO:0032010 label: phagolysosome parts: - order: 1 role: macromolecule digestion node: id: macromolecule_digestion label: Digestion of cargo macromolecules module_type: BIOLOGICAL_PROCESS description: Acid hydrolases delivered by lysosome fusion break down the cargo's DNA and proteins, and in bacteria-eating amoebae its cell wall. parts: - order: 1 role: DNA digestion node: id: cargo_dna_digestion label: DNA digestion module_type: BIOLOGICAL_PROCESS concepts: - preferred_term: DNA catabolic process term: id: GO:0006308 label: DNA catabolic process annotons: - id: dnase2_digestion label: DNase II digests engulfed DNA participant: selector_type: FAMILY family: preferred_term: DNase II term: id: PANTHER:PTHR10858 label: DEOXYRIBONUCLEASE II representative_members: - preferred_term: DNASE2 (human) term: id: UniProtKB:O00115 label: Deoxyribonuclease-2-alpha - preferred_term: Dnase2 (mouse) term: id: UniProtKB:P56542 label: Deoxyribonuclease-2-alpha - preferred_term: nuc-1 (C. elegans) term: id: UniProtKB:Q17778 label: Deoxyribonuclease-2 - preferred_term: DNaseII (D. melanogaster) term: id: UniProtKB:Q9VED8 label: Deoxyribonuclease II ancestral_nodes: - preferred_term: PAINT node PTN000092274 term: id: PANTHER:PTN000092274 label: PTN000092274 description: Eukaryote-level PAINT IBD node (taxon:2759) in PTHR10858 propagating F:GO:0004531 deoxyribonuclease II activity; human DNASE2 (O00115), mouse Dnase2a (MGI:MGI:1329019) and Drosophila DNaseII (FB:FBgn0000477) are among the seeds. evidence: - source_id: GO_REF:0000033 statement: IBD propagation of deoxyribonuclease II activity from PAINT node PTN000092274 in PTHR10858. function: preferred_term: deoxyribonuclease II activity term: id: GO:0004531 label: deoxyribonuclease II activity processes: - preferred_term: DNA catabolic process term: id: GO:0006308 label: DNA catabolic process locations: - preferred_term: phagolysosome term: id: GO:0032010 label: phagolysosome role_description: Acid DNase that finishes digesting the DNA of engulfed cells in the phagolysosome. Loss leaves undigested DNA, which in mouse and fly triggers innate immune signalling. evidence: - source_id: PMID:10716942 title: NUC-1, a caenorhabditis elegans DNase II homolog, functions in an intermediate step of DNA degradation during apoptosis. statement: In C. elegans, nuc-1 completes degradation of apoptotic-cell DNA, and completion needs engulfment. supporting_text: Completion of DNA degradation did not occur in the absence of cell-corpse engulfment. - source_id: PMID:12524536 title: Impaired thymic development in mouse embryos deficient in apoptotic DNA degradation. statement: In mouse, macrophage DNase II digests the DNA of phagocytosed apoptotic cells. supporting_text: But, when apoptotic cells were phagocytosed by macrophages, their DNA was degraded by DNase II. - source_id: PMID:11375492 title: Requirement of DNase II for definitive erythropoiesis in the mouse fetal liver. statement: DNase II acts non-cell-autonomously, in the macrophages that engulf expelled erythroid nuclei. supporting_text: suggesting that DNase II functions in a non-cell-autonomous manner - source_id: PMID:12381665 title: Activation of the innate immunity in Drosophila by endogenous chromosomal DNA that escaped apoptotic degradation. statement: A Drosophila DNase II loss-of-function mutant accumulates undigested DNA. supporting_text: We also identified a loss-of-function mutant in Drosophila for DNase II-like acid DNase. notes: 'An OpenScientist phylogenetic profile (file:human/DNASE2/DNASE2-hypotheses/dnase2-phagotroph-distribution/openscientist.md) found the family ancient and broadly eukaryotic but patchy: present in Dictyostelium (about 14 uncharacterised paralogs), Acanthamoeba, ciliates and land plants, and absent from choanoflagellates, kinetoplastids and Giardia. Presence does not track phagotrophy, so the amoebal paralogs are not assumed to digest phagocytosed DNA.' - order: 2 role: protein digestion node: id: cargo_protein_digestion label: Protein digestion module_type: BIOLOGICAL_PROCESS concepts: - preferred_term: protein processing in phagocytic vesicle term: id: GO:1900756 label: protein processing in phagocytic vesicle annotons: - id: cathepsin_b label: Cathepsin B digests engulfed cells participant: selector_type: FAMILY family: preferred_term: cathepsin B (papain-like C1 cysteine protease) term: id: PANTHER:PTHR12411:SF16 label: CATHEPSIN B representative_members: - preferred_term: CTSB (human) term: id: UniProtKB:P07858 label: Cathepsin B - preferred_term: Ctsb (mouse) term: id: UniProtKB:P10605 label: Cathepsin B - preferred_term: ctsba (zebrafish) term: id: UniProtKB:Q6PH75 label: Cathepsin B function: preferred_term: cysteine-type endopeptidase activity term: id: GO:0004197 label: cysteine-type endopeptidase activity processes: - preferred_term: protein processing in phagocytic vesicle term: id: GO:1900756 label: protein processing in phagocytic vesicle locations: - preferred_term: phagolysosome term: id: GO:0032010 label: phagolysosome role_description: Lysosomal cysteine protease needed to digest engulfed neurons in microglia; mutants accumulate undigested dead cells in phagosomes. evidence: - source_id: PMID:41826292 title: Microglial cathepsin B is necessary for neuronal efferocytosis in zebrafish and mice during brain development. statement: Cathepsin B loss leaves microglia full of undigested dead cells, in zebrafish and in mice. supporting_text: Genetic disruption of cathepsin B in zebrafish and mice had an accumulation of microglia containing undigested dead cells. - id: cathepsin_d label: Cathepsin D in the phagosome participant: selector_type: FAMILY family: preferred_term: cathepsin D (A1 aspartic protease) term: id: PANTHER:PTHR47966:SF42 label: CATHEPSIN D representative_members: - preferred_term: CTSD (human) term: id: UniProtKB:P07339 label: Cathepsin D - preferred_term: Ctsd (mouse) term: id: UniProtKB:P18242 label: Cathepsin D function: preferred_term: aspartic-type endopeptidase activity term: id: GO:0004190 label: aspartic-type endopeptidase activity processes: - preferred_term: protein processing in phagocytic vesicle term: id: GO:1900756 label: protein processing in phagocytic vesicle locations: - preferred_term: phagolysosome term: id: GO:0032010 label: phagolysosome role_description: Lysosomal aspartic protease. Ctsd-deficient cells let ingested Listeria survive longer in phagosomes. evidence: - source_id: PMID:16424157 title: 'Cutting edge: a novel nonoxidative phagosomal mechanism exerted by cathepsin-D controls Listeria monocytogenes intracellular growth.' statement: Cathepsin D acts in a nonoxidative phagosomal mechanism against ingested bacteria. supporting_text: In this study, we show that the lysosomal aspartyl-protease cathepsin-D (Ctsd) is of considerable importance for nonoxidative listericidal defense mechanisms. notes: Evidence is microbial killing rather than digestion of cells; included as the aspartic-protease arm of phagosomal proteolysis. - order: 3 role: bacterial cell wall digestion optional: true node: id: bacterial_cell_wall_digestion label: Bacterial cell wall digestion module_type: BIOLOGICAL_PROCESS concepts: - preferred_term: peptidoglycan catabolic process term: id: GO:0009253 label: peptidoglycan catabolic process annotons: - id: dicty_lysozyme label: Amoebal lysozyme digests ingested bacteria participant: selector_type: GENE_PRODUCT gene_product: preferred_term: alyA (Dictyostelium discoideum) term: id: UniProtKB:Q8T1G4 label: Lysozyme A function: preferred_term: lysozyme activity term: id: GO:0003796 label: lysozyme activity processes: - preferred_term: peptidoglycan catabolic process term: id: GO:0009253 label: peptidoglycan catabolic process locations: - preferred_term: phagolysosome term: id: GO:0032010 label: phagolysosome role_description: Phagotrophic amoebae digest the cell walls of bacteria they eat. Mutants with low lysozyme grow poorly on bacteria. evidence: - source_id: PMID:15640146 title: A Dictyostelium mutant with reduced lysozyme levels compensates by increased phagocytic activity. statement: Reduced lysozyme impairs growth on bacteria as the only food. supporting_text: The mutant cells had greatly reduced enzymatic activity and grew inefficiently when bacteria were the sole food source. notes: No PANTHER family is assigned to alyA in UniProt, so it is grounded as a gene product. - order: 2 role: export of digestion products node: id: product_export label: Export of digestion products module_type: TRANSPORT_STEP description: Transporters in the phagolysosomal membrane return amino acids, nucleosides, heme and cholesterol to the cytosol. Blocking export also stalls digestion. variant_sets: - id: exported_product label: Exported product axis: digestion product exported selection: ONE_OR_MORE variants: - id: amino_acid_export label: Basic amino acid export module_type: TRANSPORT_STEP annotons: - id: laat1_amino_acid_export label: LAAT-1/PQLC2 exports basic amino acids participant: selector_type: FAMILY family: preferred_term: LAAT-1/PQLC2 PQ-loop amino acid exporter term: id: PANTHER:PTHR16201 label: SEVEN TRANSMEMBRANE PROTEIN 1-RELATED representative_members: - preferred_term: laat-1 (C. elegans) term: id: UniProtKB:Q95XZ6 label: Lysosomal amino acid transporter 1 - preferred_term: SLC66A1/PQLC2 (human) term: id: UniProtKB:Q6ZP29 label: Lysosomal amino acid transporter 1 homolog function: preferred_term: basic amino acid transmembrane transporter activity term: id: GO:0015174 label: basic amino acid transmembrane transporter activity locations: - preferred_term: phagolysosome membrane term: id: GO:0061474 label: phagolysosome membrane role_description: Exports lysine and arginine from lysosomes. laat-1 mutants engulf corpses normally but fail to degrade them in phagolysosomes. evidence: - source_id: PMID:22822152 title: LAAT-1 is the lysosomal lysine/arginine transporter that maintains amino acid homeostasis. statement: Loss of laat-1 makes lysosomes defective in degradation. supporting_text: Loss of laat-1 caused accumulation of lysine and arginine in enlarged, degradation-defective lysosomes. - source_id: PMID:22822152 title: LAAT-1 is the lysosomal lysine/arginine transporter that maintains amino acid homeostasis. statement: Engulfment is normal in laat-1 mutants, so the defect is in digestion. supporting_text: Cell death and cell corpse engulfment were normal in qx42 mutants - source_id: PMID:23169667 title: Heptahelical protein PQLC2 is a lysosomal cationic amino acid exporter underlying the action of cysteamine in cystinosis therapy. statement: Human PQLC2 is a lysosomal exporter of cationic amino acids. supporting_text: We also show that PQLC2, a mammalian PQ-loop protein closely related to yeast Ypq proteins, localizes to lysosomes and catalyzes a robust, electrogenic transport that is selective for CAAs and strongly activated at low extracytosolic pH. notes: PQLC2 is included by orthology; it has no phagocytic-cargo data. - id: nucleoside_export label: Nucleoside export module_type: TRANSPORT_STEP annotons: - id: ent3_nucleoside_export label: ENT3 exports nucleosides participant: selector_type: FAMILY family: preferred_term: ENT3 term: id: PANTHER:PTHR10332:SF17 label: EQUILIBRATIVE NUCLEOSIDE TRANSPORTER 3 representative_members: - preferred_term: SLC29A3 (human) term: id: UniProtKB:Q9BZD2 label: Equilibrative nucleoside transporter 3 - preferred_term: Slc29a3 (mouse) term: id: UniProtKB:Q99P65 label: Equilibrative nucleoside transporter 3 function: preferred_term: nucleoside transmembrane transporter activity term: id: GO:0005337 label: nucleoside transmembrane transporter activity processes: - preferred_term: nucleoside transmembrane transport term: id: GO:1901642 label: nucleoside transmembrane transport locations: - preferred_term: phagolysosome membrane term: id: GO:0061474 label: phagolysosome membrane role_description: Lysosomal nucleoside transporter. Without it, macrophages clearing apoptotic cells accumulate lysosomal nucleosides. evidence: - source_id: PMID:22174130 title: Equilibrative nucleoside transporter 3 deficiency perturbs lysosome function and macrophage homeostasis. statement: ENT3 loss causes lysosomal nucleoside build-up during apoptotic cell clearance. supporting_text: In the absence of ENT3, defective apoptotic cell clearance led to lysosomal nucleoside buildup, elevated intralysosomal pH, and altered macrophage function. - id: heme_export label: Heme export module_ref: heme_induced_iron_recycling_macrophage_differentiation#phagolysosomal_heme_export notes: Animal-specific route. An OpenScientist phylogenetic profile (file:human/SLC48A1/SLC48A1-hypotheses/hrg1-protist-distribution/openscientist.md) found the HRG1 family almost entirely in animals and choanoflagellates, with a single amoebozoan ortholog in Acanthamoeba castellanii and none in Dictyostelium discoideum, so phagotrophic protists generally lack this exporter. - id: cholesterol_export label: Cholesterol export module_type: TRANSPORT_STEP annotons: - id: npc1_cholesterol_export label: NPC1 moves cargo cholesterol out of the lysosome participant: selector_type: FAMILY family: preferred_term: NPC1 term: id: PANTHER:PTHR45727:SF2 label: NPC INTRACELLULAR CHOLESTEROL TRANSPORTER 1 representative_members: - preferred_term: NPC1 (human) term: id: UniProtKB:O15118 label: NPC intracellular cholesterol transporter 1 - preferred_term: Npc1 (mouse) term: id: UniProtKB:O35604 label: NPC intracellular cholesterol transporter 1 function: preferred_term: cholesterol transfer activity term: id: GO:0120020 label: cholesterol transfer activity processes: - preferred_term: intracellular cholesterol transport term: id: GO:0032367 label: intracellular cholesterol transport locations: - preferred_term: phagolysosome membrane term: id: GO:0061474 label: phagolysosome membrane role_description: Lysosomal cholesterol exporter. Myeloid Npc1 deletion impairs digestion of engulfed cells. evidence: - source_id: PMID:39547438 title: Deficiency of myeloid NPC1 exacerbates liver injury and fibrosis by impairing macrophage efferocytosis. statement: Npc1-deficient macrophages degrade engulfed cargo poorly. supporting_text: In vitro mechanistical studies reveal that Npc1-deficient macrophages exhibited inefficient efferocytosis, partly due to impaired cargo degradation. notes: General lysosomal machinery; one myeloid-knockout study links it to engulfed cargo. connections: - source: macromolecule_digestion target: product_export connection_type: PROVIDES_INPUT_FOR description: Hydrolysis releases the products that the transporters export. knowledge_gaps: - gap_statement: Which hydrolases digest the protein of engulfed cell corpses in C. elegans is unknown. boundary: Corpse degradation needs lysosome function (nuc-1, laat-1), but no single cathepsin has been shown to be required for corpse digestion in the worm. gap_kind: - BIOLOGY dark_aspect: BP_DARK status: OPEN significance: The protein-digestion part has no nematode exemplar. resolution: Test cathepsin (cpr, cpl, asp) mutants and combinations for persistent corpses in engulfing cells. provenance: - reference_id: PMID:22822152 supporting_text: Cell death and cell corpse engulfment were normal in qx42 mutants