TRIT1 is the human tRNA isopentenyltransferase (IPTase), the homolog of bacterial MiaA and yeast Mod5. It transfers the dimethylallyl group of dimethylallyl diphosphate onto N6 of adenosine 37, adjacent to the anticodon, forming N6-isopentenyladenosine (i6A37). Alternative splicing gives isoforms with or without an N-terminal mitochondrial targeting sequence, so TRIT1 acts both in the cytosol and in the mitochondrial matrix. Its substrate set is small - cytosolic tRNA(Ser) (AGA, CGA, UGA anticodons), partial modification of selenocysteine tRNA(Sec), and mitochondrial tRNA(Ser)(UCN) and tRNA(Trp) - and i6A37 supports translation efficiency and fidelity, selenoprotein expression and subsequent METTL8-dependent m3C32 modification of mt-tRNA(Ser)(UCN). The enzyme has a P-loop isopentenyltransferase domain that binds DMAPP and a C-terminal matrin-type zinc finger. Biallelic TRIT1 variants abolish i6A37 in cytosolic and mitochondrial tRNAs and cause combined oxidative phosphorylation deficiency 35 (developmental delay, microcephaly, seizures).
| GO Term | Evidence | Action | Reason |
|---|---|---|---|
| GO:0003676 nucleic acid binding | IEA GO_REF:0000002 | MODIFY | Summary: InterPro mapping of nucleic acid binding from the C-terminal matrin/U1-type C2H2 zinc finger. Reason: The nucleic acid TRIT1 binds is its tRNA substrate; the anticodon stem-loop is recognised through a series of basic residues. tRNA binding (GO:0000049) is the informative, accurate term. Proposed replacements: tRNA binding Supporting Evidence: PMID:24901367 Modelling of the p.Arg323Gln mutation on the co-crystal structure of the homologous yeast isopentenyltransferase bound to a substrate tRNA, indicates that it is one of a series of adjacent basic side chains that interact with the tRNA backbone of the anticodon stem |
| GO:0005524 ATP binding | IEA GO_REF:0000002 | REMOVE | Summary: InterPro mapping of ATP binding from the P-loop NTPase fold of the isopentenyltransferase domain. Reason: tRNA isopentenyltransferases (MiaA/Mod5/TRIT1) retain a P-loop NTPase fold, but the P-loop binds the pyrophosphate of the prenyl donor dimethylallyl diphosphate (DMAPP), not ATP. TRIT1 transfers the dimethylallyl group of DMAPP to A37; no ATP-binding or ATP-dependent step is part of the reaction. The generic P-loop to ATP binding mapping is a known electronic over-reach. Supporting Evidence: file:human/TRIT1/TRIT1-uniprot.txt /ligand="dimethylallyl diphosphate" PMID:24901367 the isopentenyl group of DMAPP (dimethylallyl pyrophosphate) is transferred to N6 of A37 in substrate tRNAs by the IPTase TRIT1 |
| GO:0005737 cytoplasm | IEA GO_REF:0000044 | ACCEPT | Summary: UniProt SubCell mapping to cytoplasm (isoforms 2, 3 and 5 lacking the mitochondrial targeting sequence). Reason: TRIT1 acts in two compartments; it is detected in the cytosolic fraction and modifies cytosolic tRNA(Ser) and tRNA(Sec), as well as mitochondrial tRNAs. Supporting Evidence: PMID:24901367 TRIT1 localized with eIF4E in the cytosol (lane 2) and showed the same profile as GDH (lanes 3β6), but was undetectable in the inner mitochondrial membrane fraction |
| GO:0005739 mitochondrion | HTP PMID:34800366 Quantitative high-confidence human mitochondrial proteome an... | ACCEPT | Summary: Detected in the high-confidence human mitochondrial proteome (MitoCoP). Reason: Consistent with targeted sub-fractionation data. Supporting Evidence: PMID:34800366 mitochondrial high-confidence proteome of >1,100 proteins (MitoCoP). |
| GO:0005739 mitochondrion | IBA GO_REF:0000033 | ACCEPT | Summary: Mitochondrial localization (IBA, HPA immunofluorescence, sub-fractionation IDA, SubCell IEA for isoforms 1 and 4 with an N-terminal transit peptide). Reason: TRIT1 is present in proteinase K-protected mitoplasts and modifies mitochondrial tRNAs; core location for its mitochondrial function. Supporting Evidence: PMID:24901367 TRIT1 was present in the cytosolic fraction (Figure 2C, lane 2) and also detectable in proteinase K-treated mitoplasts (lane 6), consistent with the enzyme being present in the mitochondrial matrix |
| GO:0005739 mitochondrion | IDA GO_REF:0000052 | ACCEPT | Summary: Mitochondrial localization (IBA, HPA immunofluorescence, sub-fractionation IDA, SubCell IEA for isoforms 1 and 4 with an N-terminal transit peptide). Reason: TRIT1 is present in proteinase K-protected mitoplasts and modifies mitochondrial tRNAs; core location for its mitochondrial function. Supporting Evidence: PMID:24901367 TRIT1 was present in the cytosolic fraction (Figure 2C, lane 2) and also detectable in proteinase K-treated mitoplasts (lane 6), consistent with the enzyme being present in the mitochondrial matrix |
| GO:0005739 mitochondrion | IDA PMID:24901367 Defective i6A37 modification of mitochondrial and cytosolic ... | ACCEPT | Summary: Mitochondrial localization (IBA, HPA immunofluorescence, sub-fractionation IDA, SubCell IEA for isoforms 1 and 4 with an N-terminal transit peptide). Reason: TRIT1 is present in proteinase K-protected mitoplasts and modifies mitochondrial tRNAs; core location for its mitochondrial function. Supporting Evidence: PMID:24901367 TRIT1 was present in the cytosolic fraction (Figure 2C, lane 2) and also detectable in proteinase K-treated mitoplasts (lane 6), consistent with the enzyme being present in the mitochondrial matrix |
| GO:0005739 mitochondrion | IEA GO_REF:0000044 | ACCEPT | Summary: Mitochondrial localization (IBA, HPA immunofluorescence, sub-fractionation IDA, SubCell IEA for isoforms 1 and 4 with an N-terminal transit peptide). Reason: TRIT1 is present in proteinase K-protected mitoplasts and modifies mitochondrial tRNAs; core location for its mitochondrial function. Supporting Evidence: PMID:24901367 TRIT1 was present in the cytosolic fraction (Figure 2C, lane 2) and also detectable in proteinase K-treated mitoplasts (lane 6), consistent with the enzyme being present in the mitochondrial matrix |
| GO:0005759 mitochondrial matrix | TAS Reactome:R-HSA-6787567 | ACCEPT | Summary: Reactome places TRIT1-mediated i6A37 formation on mt-tRNA(Ser) in the mitochondrial matrix. Reason: Supported by protease protection of TRIT1 in mitoplasts and co-fractionation with the matrix marker GDH. Supporting Evidence: PMID:24901367 TRIT1 was present in the cytosolic fraction (Figure 2C, lane 2) and also detectable in proteinase K-treated mitoplasts (lane 6), consistent with the enzyme being present in the mitochondrial matrix |
| GO:0006400 tRNA modification | IBA GO_REF:0000033 | ACCEPT | Summary: TRIT1 forms N6-isopentenyladenosine at position 37 (i6A37) in a restricted set of cytosolic tRNAs (Ser-AGA/CGA/UGA, partially Sec-UCA) and mitochondrial tRNAs (Ser-UCN, Trp). Reason: Core process, supported by knockdown (IDA), patient-cell deficiency with rescue, and phylogeny (IBA). No more specific GO process term for i6A37 formation exists. Supporting Evidence: PMID:24126054 only tRNA(Ser)AGA, tRNA(Ser)CGA, tRNA(Ser)UGA, and selenocysteine tRNA with UCA |
| GO:0006400 tRNA modification | IDA PMID:24126054 Human cells have a limited set of tRNA anticodon loop substr... | ACCEPT | Summary: TRIT1 forms N6-isopentenyladenosine at position 37 (i6A37) in a restricted set of cytosolic tRNAs (Ser-AGA/CGA/UGA, partially Sec-UCA) and mitochondrial tRNAs (Ser-UCN, Trp). Reason: Core process, supported by knockdown (IDA), patient-cell deficiency with rescue, and phylogeny (IBA). No more specific GO process term for i6A37 formation exists. Supporting Evidence: PMID:24126054 only tRNA(Ser)AGA, tRNA(Ser)CGA, tRNA(Ser)UGA, and selenocysteine tRNA with UCA |
| GO:0006400 tRNA modification | IEA GO_REF:0000120 | ACCEPT | Summary: TRIT1 forms N6-isopentenyladenosine at position 37 (i6A37) in a restricted set of cytosolic tRNAs (Ser-AGA/CGA/UGA, partially Sec-UCA) and mitochondrial tRNAs (Ser-UCN, Trp). Reason: Core process, supported by knockdown (IDA), patient-cell deficiency with rescue, and phylogeny (IBA). No more specific GO process term for i6A37 formation exists. Supporting Evidence: PMID:24126054 only tRNA(Ser)AGA, tRNA(Ser)CGA, tRNA(Ser)UGA, and selenocysteine tRNA with UCA |
| GO:0008033 tRNA processing | IEA GO_REF:0000002 | ACCEPT | Summary: InterPro mapping to tRNA processing. Reason: Correct general parent of tRNA modification. |
| GO:0008270 zinc ion binding | IEA GO_REF:0000002 | KEEP AS NON CORE | Summary: InterPro mapping of zinc ion binding from the C-terminal matrin-type C2H2 zinc finger. Reason: The matrin-type zinc finger is predicted to coordinate zinc, but zinc binding has not been shown for human TRIT1 and is a structural feature of a substrate-recognition domain rather than the enzyme's function. The zinc-finger motif was noted when the human gene was cloned and is absent from prokaryotic homologues. Supporting Evidence: PMID:11111046 The human sequence was found to contain a single C2H2 Zn-finger-like motif |
| GO:0052381 tRNA dimethylallyltransferase activity | EXP PMID:24901367 Defective i6A37 modification of mitochondrial and cytosolic ... | ACCEPT | Summary: TRIT1 transfers the dimethylallyl group of DMAPP to N6 of A37 in substrate tRNAs (EC 2.5.1.75); shown by yeast mod5 complementation, i6A37 loss on knockdown and in patient cells, and in vitro modification by recombinant TRIT1. Reason: Core molecular function supported by multiple independent experimental lines and by phylogeny. Supporting Evidence: PMID:24901367 the isopentenyl group of DMAPP (dimethylallyl pyrophosphate) is transferred to N6 of A37 in substrate tRNAs by the IPTase TRIT1 PMID:11111046 Expression of the gene in a Saccharomyces cerevisiae mutant lacking the endogenous tRNA isopentenyl transferase MOD5 resulted in functional complementation and reintroduction of isopentenyladenosine into tRNA PMID:34774131 N6-isopenteny-ladenosine was introduce in 1 ΞΌM mt-tRNA by 250 nM TRIT1 and 10 ΞΌM Ξ³,Ξ³-Dimethylallyl pyrophosphate (DMAPP) |
| GO:0052381 tRNA dimethylallyltransferase activity | IBA GO_REF:0000033 | ACCEPT | Summary: TRIT1 transfers the dimethylallyl group of DMAPP to N6 of A37 in substrate tRNAs (EC 2.5.1.75); shown by yeast mod5 complementation, i6A37 loss on knockdown and in patient cells, and in vitro modification by recombinant TRIT1. Reason: Core molecular function supported by multiple independent experimental lines and by phylogeny. Supporting Evidence: PMID:24901367 the isopentenyl group of DMAPP (dimethylallyl pyrophosphate) is transferred to N6 of A37 in substrate tRNAs by the IPTase TRIT1 PMID:11111046 Expression of the gene in a Saccharomyces cerevisiae mutant lacking the endogenous tRNA isopentenyl transferase MOD5 resulted in functional complementation and reintroduction of isopentenyladenosine into tRNA PMID:34774131 N6-isopenteny-ladenosine was introduce in 1 ΞΌM mt-tRNA by 250 nM TRIT1 and 10 ΞΌM Ξ³,Ξ³-Dimethylallyl pyrophosphate (DMAPP) |
| GO:0052381 tRNA dimethylallyltransferase activity | IDA PMID:24126054 Human cells have a limited set of tRNA anticodon loop substr... | ACCEPT | Summary: TRIT1 transfers the dimethylallyl group of DMAPP to N6 of A37 in substrate tRNAs (EC 2.5.1.75); shown by yeast mod5 complementation, i6A37 loss on knockdown and in patient cells, and in vitro modification by recombinant TRIT1. Reason: Core molecular function supported by multiple independent experimental lines and by phylogeny. Supporting Evidence: PMID:24901367 the isopentenyl group of DMAPP (dimethylallyl pyrophosphate) is transferred to N6 of A37 in substrate tRNAs by the IPTase TRIT1 PMID:11111046 Expression of the gene in a Saccharomyces cerevisiae mutant lacking the endogenous tRNA isopentenyl transferase MOD5 resulted in functional complementation and reintroduction of isopentenyladenosine into tRNA PMID:34774131 N6-isopenteny-ladenosine was introduce in 1 ΞΌM mt-tRNA by 250 nM TRIT1 and 10 ΞΌM Ξ³,Ξ³-Dimethylallyl pyrophosphate (DMAPP) |
| GO:0052381 tRNA dimethylallyltransferase activity | IDA PMID:34774131 Balancing of mitochondrial translation through METTL8-mediat... | ACCEPT | Summary: TRIT1 transfers the dimethylallyl group of DMAPP to N6 of A37 in substrate tRNAs (EC 2.5.1.75); shown by yeast mod5 complementation, i6A37 loss on knockdown and in patient cells, and in vitro modification by recombinant TRIT1. Reason: Core molecular function supported by multiple independent experimental lines and by phylogeny. Supporting Evidence: PMID:24901367 the isopentenyl group of DMAPP (dimethylallyl pyrophosphate) is transferred to N6 of A37 in substrate tRNAs by the IPTase TRIT1 PMID:11111046 Expression of the gene in a Saccharomyces cerevisiae mutant lacking the endogenous tRNA isopentenyl transferase MOD5 resulted in functional complementation and reintroduction of isopentenyladenosine into tRNA PMID:34774131 N6-isopenteny-ladenosine was introduce in 1 ΞΌM mt-tRNA by 250 nM TRIT1 and 10 ΞΌM Ξ³,Ξ³-Dimethylallyl pyrophosphate (DMAPP) |
| GO:0052381 tRNA dimethylallyltransferase activity | IEA GO_REF:0000120 | ACCEPT | Summary: TRIT1 transfers the dimethylallyl group of DMAPP to N6 of A37 in substrate tRNAs (EC 2.5.1.75); shown by yeast mod5 complementation, i6A37 loss on knockdown and in patient cells, and in vitro modification by recombinant TRIT1. Reason: Core molecular function supported by multiple independent experimental lines and by phylogeny. Supporting Evidence: PMID:24901367 the isopentenyl group of DMAPP (dimethylallyl pyrophosphate) is transferred to N6 of A37 in substrate tRNAs by the IPTase TRIT1 PMID:11111046 Expression of the gene in a Saccharomyces cerevisiae mutant lacking the endogenous tRNA isopentenyl transferase MOD5 resulted in functional complementation and reintroduction of isopentenyladenosine into tRNA PMID:34774131 N6-isopenteny-ladenosine was introduce in 1 ΞΌM mt-tRNA by 250 nM TRIT1 and 10 ΞΌM Ξ³,Ξ³-Dimethylallyl pyrophosphate (DMAPP) |
| GO:0052381 tRNA dimethylallyltransferase activity | TAS PMID:11111046 Cloning of a human tRNA isopentenyl transferase. | ACCEPT | Summary: TRIT1 transfers the dimethylallyl group of DMAPP to N6 of A37 in substrate tRNAs (EC 2.5.1.75); shown by yeast mod5 complementation, i6A37 loss on knockdown and in patient cells, and in vitro modification by recombinant TRIT1. Reason: Core molecular function supported by multiple independent experimental lines and by phylogeny. Supporting Evidence: PMID:24901367 the isopentenyl group of DMAPP (dimethylallyl pyrophosphate) is transferred to N6 of A37 in substrate tRNAs by the IPTase TRIT1 PMID:11111046 Expression of the gene in a Saccharomyces cerevisiae mutant lacking the endogenous tRNA isopentenyl transferase MOD5 resulted in functional complementation and reintroduction of isopentenyladenosine into tRNA PMID:34774131 N6-isopenteny-ladenosine was introduce in 1 ΞΌM mt-tRNA by 250 nM TRIT1 and 10 ΞΌM Ξ³,Ξ³-Dimethylallyl pyrophosphate (DMAPP) |
| GO:0070900 mitochondrial tRNA modification | IDA PMID:24901367 Defective i6A37 modification of mitochondrial and cytosolic ... | ACCEPT | Summary: TRIT1 forms i6A37 on mitochondrial tRNA(Ser)(UCN) and tRNA(Trp); patient cells with TRIT1 p.Arg323Gln lack mitochondrial i6A37, rescued by wild-type TRIT1, and TRIT1-modified mt-tRNA(Ser) becomes a substrate for METTL8. Reason: Core process for the mitochondrial isoform, with direct patient-cell and in vitro evidence. Supporting Evidence: PMID:24901367 Complete complementation of the i6A37 deficiency of both cytosolic and mitochondrial tRNAs was achieved by transduction of patient fibroblasts with wild-type TRIT1 PMID:34774131 our biochemical reconstitution revealed that METTL8 requires prior TRIT1 activity, presumably to generate the i6A37 modification to set the m3C32 mark on the mt-tRNASer(UCN) |
| GO:0070900 mitochondrial tRNA modification | IDA PMID:34774131 Balancing of mitochondrial translation through METTL8-mediat... | ACCEPT | Summary: TRIT1 forms i6A37 on mitochondrial tRNA(Ser)(UCN) and tRNA(Trp); patient cells with TRIT1 p.Arg323Gln lack mitochondrial i6A37, rescued by wild-type TRIT1, and TRIT1-modified mt-tRNA(Ser) becomes a substrate for METTL8. Reason: Core process for the mitochondrial isoform, with direct patient-cell and in vitro evidence. Supporting Evidence: PMID:24901367 Complete complementation of the i6A37 deficiency of both cytosolic and mitochondrial tRNAs was achieved by transduction of patient fibroblasts with wild-type TRIT1 PMID:34774131 our biochemical reconstitution revealed that METTL8 requires prior TRIT1 activity, presumably to generate the i6A37 modification to set the m3C32 mark on the mt-tRNASer(UCN) |
| GO:0070900 mitochondrial tRNA modification | TAS Reactome:R-HSA-6787567 | ACCEPT | Summary: TRIT1 forms i6A37 on mitochondrial tRNA(Ser)(UCN) and tRNA(Trp); patient cells with TRIT1 p.Arg323Gln lack mitochondrial i6A37, rescued by wild-type TRIT1, and TRIT1-modified mt-tRNA(Ser) becomes a substrate for METTL8. Reason: Core process for the mitochondrial isoform, with direct patient-cell and in vitro evidence. Supporting Evidence: PMID:24901367 Complete complementation of the i6A37 deficiency of both cytosolic and mitochondrial tRNAs was achieved by transduction of patient fibroblasts with wild-type TRIT1 PMID:34774131 our biochemical reconstitution revealed that METTL8 requires prior TRIT1 activity, presumably to generate the i6A37 modification to set the m3C32 mark on the mt-tRNASer(UCN) |
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Download this section (compressed HTML)Q: What restricts human TRIT1 to so few A36-A37-A38 tRNAs compared with the yeast enzymes, and does the C-terminal zinc finger contribute to substrate selection?
Q: Do the cytosolic and mitochondrial TRIT1 pools contribute differently to COXPD35 pathology?
Experiment: Compartment-restricted rescue of TRIT1-null cells (mitochondrially targeted versus cytosol-only TRIT1) followed by i6A37 mapping and mitochondrial translation assays.
Experiment: In vitro binding and activity assays of TRIT1 lacking the zinc finger, with substrate and non-substrate A36-A38 tRNAs.
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