AARS2 is a nuclear-encoded, monomeric class II alanyl-tRNA synthetase that is imported into mitochondria. It activates L-alanine with ATP and transfers alanine to mitochondrial tRNA(Ala), supplying Ala-tRNA(Ala) for mitochondrial translation. Its editing apparatus hydrolyzes mischarged Ser-tRNA(Ala) and Gly-tRNA(Ala), thereby preserving mitochondrial translational fidelity, while its C-Ala domain contributes to recognition of the structurally idiosyncratic mitochondrial tRNA(Ala). Under hypoxia or elevated intracellular lactate, AARS2 can additionally use ATP and lactate to lactylate protein lysines on mitochondrial substrates such as PDHA1 and CPT2. Lactate-responsive cGAS lactylation has also been reported, although how mitochondrial AARS2 accesses cytosolic or nuclear cGAS remains unresolved. Pathogenic biallelic variants can impair AARS2 abundance or alanine charging and cause tissue-selective mitochondrial disease, including infantile cardiomyopathy and leukodystrophy with ovarian failure.
Definition: A peptidyl-lysine modification process in which a lactyl group is covalently added to the epsilon-amino group of a lysine residue within a protein.
Justification: GO:0141207 captures the ATP-dependent molecular activity but no current biological process term captures its immediate protein-modification outcome. The proposed term should be placed under GO:0018205 peptidyl-lysine modification and related to the broader GO:0043543 protein acylation branch; it would support curation of AARS2 and other experimentally established lysine lactylation mechanisms without conflating them with downstream signaling phenotypes. A live QuickGO ontology search on 2026-09-25 found related delactylase activities but no peptidyl-lysine lactylation biological-process term.
Parent term: peptidyl-lysine modification
Supporting Evidence:
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0005739 mitochondrion | IBA GO_REF:0000033 | ACCEPT | Summary: IBA assignment of AARS2 activity in mitochondria, independently supported by mitochondrial import, enzymology, proteomics, and hypoxia-response studies. Reason: Mitochondria are the established execution site for AARS2's canonical tRNA aminoacylation and editing functions and for its hypoxia-responsive lactylation of mitochondrial substrates. Propagation Review Root cause: NO FAILURE CORE Sources checked: PANTHER:PTN002622939 · PTN002622939 SUPPORTS TRANSFER PAINT ancestral node for mitochondrial localization. Direct human AARS2-GFP targeting in PMID:21549344 supports the organellar placement. The target itself among descendant evidence is expected and is not circular. |
| GO:0070143 mitochondrial alanyl-tRNA aminoacylation | IBA GO_REF:0000033 | ACCEPT | Summary: IBA assignment of the mitochondrial alanine-charging process, corroborated by direct recombinant-enzyme assays and rescue of defective mt-tRNA(Ala) charging in human cybrids. Reason: This is the organelle-specific core process executed by AARS2 and is at the appropriate GO granularity. Propagation Review Root cause: NO FAILURE CORE Sources checked: PANTHER:PTN002622939 · PTN002622939 SUPPORTS TRANSFER PAINT ancestral node for mitochondrial alanine charging. Human cybrid rescue and purified human enzyme studies corroborate this inherited process; target-derived experimental evidence is legitimate support for the ancestral assertion. Supporting Evidence: PMID:30262995 This indicated that AARS2 improved the aminoacylation activity in the case of m.5655A>G, rather than having a stabilizing effect on the tRNA structure. |
| GO:0004813 alanine-tRNA ligase activity | IBA GO_REF:0000033 | ACCEPT | Summary: IBA assignment of substrate-specific alanine-tRNA ligase activity, directly confirmed for purified human mitochondrial AlaRS and in human-cell tRNA-charging rescue experiments. Reason: Alanine-tRNA ligase activity is AARS2's principal conserved molecular function. Propagation Review Root cause: NO FAILURE CORE Sources checked: PANTHER:PTN000206950 · PTN000206950 SUPPORTS TRANSFER PAINT ancestral node for alanine-tRNA ligase activity. Direct human enzyme assays agree with the conserved catalytic assignment; the inclusion of human AARS2 among descendants is not circular propagation. Supporting Evidence: PMID:30262995 This indicated that AARS2 improved the aminoacylation activity in the case of m.5655A>G, rather than having a stabilizing effect on the tRNA structure. |
| GO:0002161 aminoacyl-tRNA deacylase activity | IBA GO_REF:0000033 | MODIFY | Summary: The conserved aminoacyl-tRNA proofreading activity is correct; direct human assays identify both mischarged Ser-tRNA(Ala) and Gly-tRNA(Ala) as substrates. Reason: Retain the biological conclusion of the PAINT inference, but refine the human annotation to Ser-tRNA(Ala) deacylase activity and Gly-tRNA(Ala) deacylase activity. PMID:29228266 and PMID:30952159 directly assay the two post-transfer reactions with human AARS2. Both terms are children of GO:0002161 in live QuickGO (2026-09-25). Recording them here as replacements avoids appending redundant NEW descendants of the retained source annotation; this is a specificity recommendation, not evidence of incorrect phylogenetic transfer. Propagation Review Root cause: TERM SCOPING PROBLEM Failure modes: GRANULARITY MISMATCH Sources checked: PANTHER:PTN000206950 · PTN000206950 SUPPORTS TRANSFER PAINT ancestral node for aminoacyl-tRNA proofreading. Direct human Ser-tRNA(Ala) and Gly-tRNA(Ala) hydrolysis supports the inherited parent activity. Proposed substrate-specific replacements refine the human endpoint without alleging a faulty ancestral placement. Proposed replacements: Ser-tRNA(Ala) deacylase activity Gly-tRNA(Ala) deacylase activity Supporting Evidence: PMID:29228266 We demonstrate here that the human mitochondrial AlaRS is capable of editing mischarged tRNAs in vitro PMID:30952159 Indeed, the result of the post-transfer editing assay showed that hmtAlaRS-R663E was unable to hydrolyze Gly-tRNAAla (Figure 5F). |
| GO:0000166 nucleotide binding | IEA GO_REF:0000002 | MODIFY | Summary: InterPro inference of generic nucleotide binding from conserved AlaRS domains; the physiologically relevant nucleotide substrate is ATP. Reason: Nucleotide binding is true but too broad. Human AARS2 uses ATP for alanine activation and for its conditional lactyltransferase reaction, so ATP binding is the informative replacement. Propagation Review Root cause: TERM SCOPING PROBLEM Failure modes: GRANULARITY MISMATCH Sources checked: InterPro:IPR002318 SUPPORTS TRANSFER The AlaRS family mapping explicitly supplies nucleotide binding. ATP-dependent alanine activation makes ATP binding the useful target-specific refinement. InterPro:IPR018162 SUPPORTS TRANSFER The class IIc domain superfamily carries nucleotide binding and ATP binding. The former is true but less specific for AARS2. InterPro:IPR018163 SUPPORTS TRANSFER The putative editing-domain superfamily supplies generic nucleotide binding; this does not specify ATP by itself. Whole-enzyme human biochemistry supplies the proposed refinement. InterPro:IPR018164 SUPPORTS TRANSFER The N-terminal AlaRS domain maps to nucleotide binding and ATP binding; biochemical ATP use supports the more informative child. Proposed replacements: ATP binding |
| GO:0003676 nucleic acid binding | IEA GO_REF:0000002 | MODIFY | Summary: InterPro inference of generic nucleic-acid binding; AARS2's established physiological nucleic-acid ligand is mitochondrial tRNA(Ala). Reason: Replace the uninformative parent with tRNA binding, which is directly supported by biochemical dissection of human mitochondrial AlaRS recognition of mt-tRNA(Ala). Propagation Review Root cause: TERM SCOPING PROBLEM Failure modes: GRANULARITY MISMATCH Sources checked: InterPro:IPR018165 SUPPORTS TRANSFER The AlaRS core-domain mapping supplies nucleic acid binding. Human mt-tRNA(Ala) recognition assays establish tRNA as the relevant ligand and support the proposed tRNA-binding refinement. Proposed replacements: tRNA binding |
| GO:0004812 aminoacyl-tRNA ligase activity | IEA GO_REF:0000002 | MODIFY | Summary: InterPro inference of broad aminoacyl-tRNA ligase activity from an aaRS additional domain. Reason: The activity class is correct, but alanine specificity is firmly established for AARS2 and should be stated explicitly. Propagation Review Root cause: TERM SCOPING PROBLEM Failure modes: GRANULARITY MISMATCH Sources checked: InterPro:IPR012947 SUPPORTS TRANSFER The threonyl/alanyl tRNA synthetase SAD domain maps to the broad ligase class. AARS2 biochemistry establishes alanine specificity, which this shared-domain mapping does not resolve. Proposed replacements: alanine-tRNA ligase activity |
| GO:0004813 alanine-tRNA ligase activity | IEA GO_REF:0000120 | ACCEPT | Summary: Combined UniRule, InterPro, EC 6.1.1.7, and RHEA:12540 assignment of alanine-tRNA ligase activity. Reason: The electronic mapping exactly matches the directly characterized core reaction of human mitochondrial AlaRS. Propagation Review Root cause: NO FAILURE CORE Sources checked: InterPro:IPR018162 SUPPORTS TRANSFER This InterPro AlaRS domain or family explicitly maps to alanine-tRNA ligase activity, concordant with direct human enzyme and cybrid evidence. InterPro:IPR018164 SUPPORTS TRANSFER This InterPro AlaRS domain or family explicitly maps to alanine-tRNA ligase activity, concordant with direct human enzyme and cybrid evidence. InterPro:IPR018165 SUPPORTS TRANSFER This InterPro AlaRS domain or family explicitly maps to alanine-tRNA ligase activity, concordant with direct human enzyme and cybrid evidence. InterPro:IPR023033 SUPPORTS TRANSFER This InterPro AlaRS domain or family explicitly maps to alanine-tRNA ligase activity, concordant with direct human enzyme and cybrid evidence. RHEA:12540 SUPPORTS TRANSFER The alanine-tRNA charging reaction matches the characterized AARS2 reaction. EC:6.1.1.7 SUPPORTS TRANSFER The substrate-specific alanine-tRNA ligase EC mapping matches the conserved human reaction. UniRule:UR000375824 SUPPORTS TRANSFER The eukaryotic AlaRS rule agrees with human aminoacylation assays; the rule is electronic support, not a separate experiment. |
| GO:0005524 ATP binding | IEA GO_REF:0000120 | ACCEPT | Summary: Electronic ATP-binding assignment from conserved AlaRS catalytic domains and UniRule. Reason: ATP is required to form the aminoacyl-adenylate intermediate in alanine charging and is also required by the conditional lactyltransferase reaction. Propagation Review Root cause: NO FAILURE CORE Sources checked: InterPro:IPR012947 SUPPORTS TRANSFER This InterPro AlaRS-associated domain explicitly carries ATP binding, consistent with ATP-dependent alanine activation by the human enzyme. InterPro:IPR018162 SUPPORTS TRANSFER This InterPro AlaRS-associated domain explicitly carries ATP binding, consistent with ATP-dependent alanine activation by the human enzyme. InterPro:IPR018164 SUPPORTS TRANSFER This InterPro AlaRS-associated domain explicitly carries ATP binding, consistent with ATP-dependent alanine activation by the human enzyme. InterPro:IPR018165 SUPPORTS TRANSFER This InterPro AlaRS-associated domain explicitly carries ATP binding, consistent with ATP-dependent alanine activation by the human enzyme. UniRule:UR000375824 SUPPORTS TRANSFER The eukaryotic AlaRS rule transfers the ATP-binding feature required for aminoacyl-adenylate synthesis. |
| GO:0005737 cytoplasm | IEA GO_REF:0000002 | MODIFY | Summary: InterPro assigns the broad cellular-component term cytoplasm from AlaRS domains. Because GO cytoplasm includes organelles, the row is not false, but it is needlessly broad for this mitochondrial enzyme. Reason: Prefer mitochondrion, the experimentally established location of mature human AARS2. Cytoplasm includes mitochondria and is therefore not contradicted. PMID:38163844 detects a hypoxia-associated cytosolic pool in mouse cells; this is not direct evidence of routine human cytosolic residence. PMID:39322678 reports human AARS2-cGAS proximity and lactylation, but does not establish which imported or precursor AARS2 species accesses cGAS. Propagation Review Root cause: TERM SCOPING PROBLEM Failure modes: GRANULARITY MISMATCH Sources checked: InterPro:IPR002318 SUPPORTS TRANSFER This AlaRS family/domain maps to cytoplasm, a term that includes mitochondria. The mapping is true but lacks the mitochondrial specificity established for human AARS2. InterPro:IPR018162 SUPPORTS TRANSFER This AlaRS family/domain maps to cytoplasm, a term that includes mitochondria. The mapping is true but lacks the mitochondrial specificity established for human AARS2. Proposed replacements: mitochondrion |
| GO:0005739 mitochondrion | IEA GO_REF:0000044 | ACCEPT | Summary: UniProt subcellular-location mapping places AARS2 in mitochondria, consistent with its targeting sequence and extensive direct human evidence. Reason: Mitochondrion is the established core compartment for AARS2 aminoacylation, editing, and hypoxia-responsive lactylation of mitochondrial substrates. Propagation Review Root cause: NO FAILURE CORE Sources checked: UniProtKB-SubCell:SL-0173 SUPPORTS TRANSFER The reviewed mitochondrial location maps directly to GO:0005739 and is independently supported by AARS2-GFP mitochondrial targeting in human HEK293T cells. |
| GO:0006419 alanyl-tRNA aminoacylation | IEA GO_REF:0000120 | MODIFY | Summary: Electronic assignment of the general alanyl-tRNA aminoacylation process; AARS2 performs this reaction specifically in mitochondria. Reason: The parent process is correct, but the mitochondrial child is both available and directly established for AARS2. Propagation Review Root cause: TERM SCOPING PROBLEM Failure modes: GRANULARITY MISMATCH Sources checked: InterPro:IPR018162 SUPPORTS TRANSFER This AlaRS-domain mapping carries the correct general alanine-charging process; mitochondrial specificity comes from the characterized human AARS2 location and substrate. InterPro:IPR018164 SUPPORTS TRANSFER This AlaRS-domain mapping carries the correct general alanine-charging process; mitochondrial specificity comes from the characterized human AARS2 location and substrate. InterPro:IPR018165 SUPPORTS TRANSFER This AlaRS-domain mapping carries the correct general alanine-charging process; mitochondrial specificity comes from the characterized human AARS2 location and substrate. ARBA:ARBA00085735 SUPPORTS TRANSFER The supplied ARBA mapping asserts alanyl-tRNA aminoacylation. Its target endpoint is correct but less specific than the directly established mitochondrial process. Proposed replacements: mitochondrial alanyl-tRNA aminoacylation |
| GO:0008270 zinc ion binding | IEA GO_REF:0000104 | ACCEPT | Summary: UniRule transfer of zinc-ion binding to the conserved AARS2 editing domain. Reason: The AlaRS editing site has conserved zinc-coordinating residues. Human C749A mutagenesis impairs deacylation, while structural modeling uses homologous editing-domain templates. Together these support the electronic zinc-binding assignment without presenting the modeling as a direct human zinc-binding assay. Propagation Review Root cause: NO FAILURE CORE Sources checked: UniRule:UR000375824 SUPPORTS TRANSFER The eukaryotic AlaRS rule transfers zinc binding. Conserved editing-site residues and human C749A deacylation assays in PMID:29228266 corroborate the zinc-dependent editing architecture; these are not direct zinc-occupancy measurements. Supporting Evidence: PMID:29228266 This cysteine is fully conserved in AlaRSs, and is critical for zinc binding in the editing active site and thus for the deacylation reaction |
| GO:0043039 tRNA aminoacylation | IEA GO_REF:0000002 | MODIFY | Summary: InterPro inference of broad tRNA aminoacylation from a conserved aminoacyl-tRNA synthetase domain. Reason: AARS2 has established alanine specificity and acts on mitochondrial tRNA(Ala), so the organelle- and substrate-specific child process is preferable. Propagation Review Root cause: TERM SCOPING PROBLEM Failure modes: GRANULARITY MISMATCH Sources checked: InterPro:IPR012947 SUPPORTS TRANSFER The shared threonyl/alanyl SAD-domain mapping supplies general tRNA aminoacylation. Direct human enzyme studies justify the mitochondrial alanine-specific replacement. Proposed replacements: mitochondrial alanyl-tRNA aminoacylation |
| GO:0106074 aminoacyl-tRNA metabolism involved in translational fidelity | IEA GO_REF:0000108 | ACCEPT | Summary: Logical inference of aminoacyl-tRNA quality-control metabolism from AARS2's editing activity. Reason: The term accurately captures removal or prevention of incorrect amino acids on mitochondrial tRNA(Ala), a core translational-fidelity function directly demonstrated for human AARS2. Propagation Review Root cause: NO FAILURE CORE Sources checked: GO:0002161 SUPPORTS TRANSFER The logical MF-to-BP inference starts from hydrolysis of incorrectly aminoacylated tRNA. Both human Ser-tRNA(Ala) and Gly-tRNA(Ala) editing assays support its participation in translational quality control. Supporting Evidence: PMID:29228266 These results indicate that tRNA proofreading is essential in mammalian mitochondria, and cannot be overcome by other quality control mechanisms. |
| GO:0005515 protein binding | IPI PMID:32296183 A reference map of the human binary protein interactome. | REMOVE | Summary: The HuRI binary-interactome screen recovered AARS2 interactions with EHHADH and PTPA isoform 2, but generic protein binding conveys no molecular role. Reason: Remove the generic protein-binding annotation because the HuRI binary interactions do not define an informative molecular role for AARS2. This is not a claim that the two interactions are false or that the experimental evidence is misattributed. The study itself states that most individual pairwise interactions lack functional characterization; neither partner supports a specific adapter, complex, or catalytic activity for AARS2. Supporting Evidence: PMID:32296183 While HuRI displays highly significant overlap with known functional relationships, the cellular function of most individual PPIs remains to be elucidated. |
| GO:0141207 peptide lactyltransferase (ATP-dependent) activity | IEA GO_REF:0000120 | ACCEPT | Summary: Electronic assignment of ATP-dependent peptide lactyltransferase activity from the exact Rhea reactions, corroborated by independent AARS2 studies of mitochondrial and cGAS substrates. Reason: AARS2 has a directly measured intrinsic ATP-dependent lactyltransferase activity. Treat it as a conditional secondary core function under elevated lactate or hypoxia and for specific substrates, not as the dominant source of basal global lysine lactylation. Propagation Review Root cause: NO FAILURE CORE Sources checked: UniProtKB:Q14CH7 · Aars2 SUPPORTS TRANSFER The reviewed mouse Aars2 record carries IDA lactyltransferase evidence. Mouse cellular studies and direct human AARS2 biochemical assays support transfer of the conditional catalytic activity. ensembl:ENSMUSP00000024733 SUPPORTS TRANSFER Mouse protein identifier supplied in the combined inference. This record is not counted as an independent experiment beyond the corresponding mouse Aars2 evidence. ARBA:ARBA00085408 SUPPORTS TRANSFER The supplied rule mapping agrees with experimentally measured ATP-dependent lactylation. It does not establish a basal global lactylome role. RHEA:80271 SUPPORTS TRANSFER ATP-dependent protein-lysine lactylation reaction underlying GO:0141207; the human catalytic experiments support this chemistry. RHEA:80275 SUPPORTS TRANSFER Additional supplied lactylation reaction mapping. Acceptance of the peptide-activity term rests on the independently measured protein-substrate reaction, not on assuming every mapped substrate is physiological. Supporting Evidence: PMID:38163844 We show that mitochondrial alanyl-tRNA synthetase (AARS2) is a protein lysine lactyltransferase |
| GO:0160049 negative regulation of cGAS/STING signaling pathway | IEA GO_REF:0000120 | KEEP AS NON CORE | Summary: AARS2-dependent cGAS lactylation inhibits cGAS/STING signaling in lactate-stimulated human and mouse experimental systems. Reason: The publisher full text of PMID:39322678 reports reduced endogenous cGAS lactylation after AARS2 depletion in human THP1 cells, together with human cell biochemical assays and mouse conditional genetics. Retain the supported negative regulatory direction as a conditional secondary process. Cytoplasmic proximity to cGAS is reported in HeLa cells, but precursor versus mature AARS2 and its import/access mechanism remain unresolved. The production GO-CAM 66f5faaa00000026 models this reaction without assigning an AARS2 compartment. Propagation Review Root cause: NO FAILURE NON CORE Sources checked: UniProtKB:Q14CH7 · Aars2 SUPPORTS TRANSFER The reviewed mouse Aars2 record carries IDA negative cGAS/STING regulation. PMID:39322678 also reports human AARS2-dependent cGAS lactylation, so the conditional pathway direction is supported across these systems. ensembl:ENSMUSP00000024733 SUPPORTS TRANSFER Mouse protein identifier supplied by the combined inference. It supports the same mouse source context rather than an independent human localization claim. ARBA:ARBA00088339 SUPPORTS TRANSFER The supplied rule maps the mouse-supported immune-regulatory outcome. Retain it as a context-dependent secondary process; it does not locate the relevant human AARS2 pool. Supporting Evidence: PMID:39322678 In response to L-lactate, AARS2 associates with cyclic GMP-AMP synthase (cGAS) and mediates its lactylation and inactivation in cells and in mice. |
| GO:0005739 mitochondrion | IDA GO_REF:0000052 | ACCEPT | Summary: Curated immunofluorescence evidence places AARS2 in mitochondria. Reason: The localization agrees with the targeting sequence, disease study, high-confidence mitochondrial proteomics, and direct mitochondrial enzymology. |
| GO:0141207 peptide lactyltransferase (ATP-dependent) activity | IDA PMID:39322678 AARS1 and AARS2 sense L-lactate to regulate cGAS as global l... | ACCEPT | Summary: Direct ATP-dependent peptide lactyltransferase annotation with cGAS as the input and negative regulation of cGAS/STING signaling as the contextual process. Reason: The paper directly supports AARS2-catalyzed lactylation, and an independent full-text study measured ATP-dependent AARS2 lactylation of mitochondrial peptide substrates. Scope this intrinsic secondary activity to elevated-lactate, hypoxic, and substrate-specific settings; AARS2 is not required for basal global Kla in HEK293T cells. How mitochondrial AARS2 accesses cGAS remains unresolved. Supporting Evidence: PMID:39322678 they directly catalyse L-lactate for ATP-dependent lactylation on the lysine acceptor end PMID:38163844 Lactylation reactions were carried out in a 30 μL reaction mix containing 50 mM pH 7.5 HEPES, 25 mM KCl, 2 mM MgCl2, 1–10 mM lactate, 4 mM ATP, and 10 nM AARS2. |
| GO:0160049 negative regulation of cGAS/STING signaling pathway | IDA PMID:39322678 AARS1 and AARS2 sense L-lactate to regulate cGAS as global l... | KEEP AS NON CORE | Summary: AARS2-dependent cGAS lactylation inhibits cGAS/STING signaling in lactate-stimulated human and mouse experimental systems. Reason: The publisher full text of PMID:39322678 reports reduced endogenous cGAS lactylation after AARS2 depletion in human THP1 cells, together with human cell biochemical assays and mouse conditional genetics. Retain the supported negative regulatory direction as a conditional secondary process. Cytoplasmic proximity to cGAS is reported in HeLa cells, but precursor versus mature AARS2 and its import/access mechanism remain unresolved. The production GO-CAM 66f5faaa00000026 models this reaction without assigning an AARS2 compartment. Supporting Evidence: PMID:39322678 In response to L-lactate, AARS2 associates with cyclic GMP-AMP synthase (cGAS) and mediates its lactylation and inactivation in cells and in mice. |
| GO:0005739 mitochondrion | HTP PMID:34800366 Quantitative high-confidence human mitochondrial proteome an... | ACCEPT | Summary: High-throughput quantitative proteomics places AARS2 in the high-confidence human mitochondrial proteome. Reason: This independent proteomic localization agrees with direct disease, enzymology, and subcellular-location evidence for AARS2's core mitochondrial residence. Supporting Evidence: PMID:34800366 We classified >8,000 proteins in mitochondrial preparations of human cells and defined a mitochondrial high-confidence proteome of >1,100 proteins (MitoCoP). |
| GO:0004813 alanine-tRNA ligase activity | IMP PMID:21549344 Exome sequencing identifies mitochondrial alanyl-tRNA synthe... | ACCEPT | Summary: The disease study linked biallelic AARS2 variants to mitochondrial respiratory-chain deficiency and proposed catalytic consequences from homology modeling. Reason: Accept the canonical alanine-charging function on the combined evidence, including later purified human AARS2 and cybrid-rescue assays. The recovered 2011 full text modeled the variant effects rather than directly measuring their aminoacylation kinetics; it also reported no mitochondrial translation defect in the cultured patient cells tested. Retain the curator-supplied IMP evidence code and original reference, but do not present those modeled defects as direct enzyme measurements. Supporting Evidence: PMID:32080176 Hs mt AlaRS charges cognate mt tRNAAla but not mt tRNAs specific for Asp, Leu, or Ser. |
| GO:0070143 mitochondrial alanyl-tRNA aminoacylation | IMP PMID:21549344 Exome sequencing identifies mitochondrial alanyl-tRNA synthe... | ACCEPT | Summary: The disease study linked biallelic AARS2 variants to mitochondrial respiratory-chain deficiency and proposed catalytic consequences from homology modeling. Reason: Accept the canonical alanine-charging function on the combined evidence, including later purified human AARS2 and cybrid-rescue assays. The recovered 2011 full text modeled the variant effects rather than directly measuring their aminoacylation kinetics; it also reported no mitochondrial translation defect in the cultured patient cells tested. Retain the curator-supplied IMP evidence code and original reference, but do not present those modeled defects as direct enzyme measurements. Supporting Evidence: PMID:32080176 Hs mt AlaRS charges cognate mt tRNAAla but not mt tRNAs specific for Asp, Leu, or Ser. |
| GO:0005739 mitochondrion | IDA PMID:21549344 Exome sequencing identifies mitochondrial alanyl-tRNA synthe... | ACCEPT | Summary: AARS2-GFP expressed in HEK293T cells was directly shown to target mitochondria in the foundational human disease study. Reason: The recovered full text reports mitochondrial targeting of the full-length human AARS2-GFP fusion in HEK293T cells (Figure 3D), directly supporting the IDA location. Later human mitochondrial proteomics independently corroborates this compartment. Supporting Evidence: PMID:21549344 AARS2-GFP fusion protein was efficiently targeted to mitochondria in the cells |
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Download this section (compressed HTML)Q: In human mitochondria, what fractions of glycine and serine errors are prevented by AARS2 pre-transfer editing versus cleared by post-transfer deacylation under physiological substrate concentrations?
Q: Across physiological and pathological lactate concentrations, which endogenous protein substrates depend materially on AARS2 rather than class I HDACs or other lactylation routes, and in which tissues and cell states?
Q: What AARS2 species and trafficking mechanism permit direct encounter with cGAS, and does this involve the mitochondrial precursor, the mature enzyme, or an inducible extramitochondrial pool?
Experiment: Introduce matched separation-of-function AARS2 variants into an acute AARS2-depletion background, then quantify charging and mischarging of mitochondrial tRNA(Ala) and Ala-to-Gly or Ala-to-Ser substitutions in pulse-labeled mitochondrial translation products. Complement the cell measurements with pre-transfer and single-turnover post-transfer kinetics using purified human AARS2.
Hypothesis: AARS2 uses mechanistically distinct proofreading routes for glycine and serine errors that make different contributions to mitochondrial translational fidelity in human cells.
Type: mitochondrial tRNA biochemistry and quantitative mitoproteomics
Experiment: Use acute endogenous AARS2 degradation across calibrated intracellular lactate and oxygen ranges, followed by isotope-resolved quantitative lactyl-proteomics and targeted measurement of PDHA1 K336 and CPT2 K457/K458. Rescue with matched-expression AARS2 variants that retain alanine charging but differ in lactyltransferase activity, and compare with class I HDAC perturbation.
Hypothesis: Endogenous AARS2 contributes substantially to a restricted, high-lactate mitochondrial lactylome but not to basal global lysine lactylation.
Type: acute perturbation and quantitative lactyl-proteomics
Experiment: Endogenously tag both termini of AARS2, distinguish precursor from mature protein, and combine quantitative fractionation, protease protection, live-cell imaging, and cGAS-directed proximity labeling before and after lactate exposure. Perturb the mitochondrial targeting sequence and import machinery while measuring site-specific cGAS lactylation and cGAS-STING signaling, with charging-competent rescue controls.
Hypothesis: A lactate-responsive AARS2 precursor or inducible extramitochondrial species directly encounters cGAS, whereas the mature mitochondrial pool does not.
Type: compartment-resolved trafficking and proximity analysis
What is not known — curated, literature-grounded statements of the open unknowns (the inverse of core functions).
Gap: The relative in vivo contributions of AARS2 pre-transfer and post-transfer editing to clearance of glycine errors, and the physiological balance between glycine and serine mischarging of mitochondrial tRNA(Ala), remain unresolved in human cells.
OPEN BIOLOGY RESIDUAL_SUBGAP
What is known: Purified human AARS2 directly edits both Gly-tRNA(Ala) and Ser-tRNA(Ala), with biochemical evidence for strong glycine pre-transfer editing and post-transfer hydrolysis of both mischarged products. These assays establish the activities but do not quantify their separate contributions under endogenous mitochondrial substrate concentrations.
Significance: Resolving the editing pathway used for each error would clarify which proofreading defect drives mitochondrial mistranslation in different AARS2 alleles and metabolic states.
What would resolve it: Use separation-of-function AARS2 variants in human cells and directly quantify aminoacylated and mischarged mitochondrial tRNA(Ala), together with Ala-to-Ser and Ala-to-Gly substitutions in newly synthesized mitochondrial proteins.
Provenance (the field's own admissions):
Gap: The physiological lactate range, cell states, tissues, and substrate spectrum in which endogenous AARS2 materially contributes to protein lysine lactylation, relative to class I HDACs and other routes, remain incompletely defined.
OPEN BIOLOGYCURATION RESIDUAL_SUBGAP
What is known: AARS2-dependent lactylation of PDHA1 and CPT2 is supported under exercise-, hypoxia-, or lactate-associated conditions, whereas AARS2 knockdown does not reduce basal global lysine lactylation in untreated HEK293T cells.
Significance: A quantitative boundary is needed to distinguish physiological AARS2 substrates from high-lactate or overexpression effects and to avoid generalizing a conditional, target-specific activity to the basal lactylome.
What would resolve it: Quantify endogenous site-resolved lactylation across calibrated intracellular lactate and oxygen ranges after acute AARS2 depletion and matched rescue, alongside class I HDAC perturbation and measurements of mitochondrial alanine charging.
Provenance (the field's own admissions):
Gap: Which AARS2 species encounters cGAS, and how its mitochondrial import or trafficking permits that encounter under elevated lactate, remain unresolved.
OPEN BIOLOGYCURATION RESIDUAL_SUBGAP
What is known: Human AARS2-GFP targets mitochondria. The cGAS study reports endogenous AARS2-cGAS proximity in human HeLa cells and AARS2-dependent cGAS lactylation in human THP1 cells, but these results do not resolve precursor versus mature enzyme or the import/access route. The production GO-CAM leaves the AARS2 activity unlocalized. A routine cytosolic or nuclear location is therefore not assigned.
Significance: Defining the relevant AARS2 species and compartment is necessary to distinguish a direct signaling mechanism from indirect metabolic effects and to curate cellular locations without over-annotation.
What would resolve it: Endogenously tag AARS2 and resolve its precursor and mature forms by quantitative fractionation, protease protection, and live-cell imaging during lactate exposure; combine this with AARS2-cGAS proximity labeling and mitochondrial-import perturbation.
Provenance (the field's own admissions):
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