A0A1S3Y076 (LOC107770656) encodes a nuclear-type proteinaceous RNase P (PRORP) of common tobacco, a single-polypeptide endonuclease that removes the 5' leader from precursor tRNAs (EC 3.1.26.5). The 540-residue protein consists of an N-terminal pentatricopeptide repeat (PPR) region that recognises the pre-tRNA body, a central zinc-binding region and a C-terminal NYN-type metallonuclease domain; all four catalytic aspartates and the four zinc ligands of characterised plant PRORPs are conserved. It belongs to the PRORP2 subfamily of the PRORP family, together with Arabidopsis PRORP2, and lacks the organellar transit peptide carried by the chloroplast and mitochondrial enzyme PRORP1. By analogy with Arabidopsis PRORP2, which localises to the nucleus, requires Mg2+ for catalysis and processes nuclear-encoded pre-tRNAs, the tobacco protein is expected to carry out 5' maturation of nuclear-encoded (cytosolic) tRNAs. In land plants protein-only enzymes of this family supply RNase P activity in the nucleus and in the organelles, in place of the ribonucleoprotein RNase P found in most other eukaryotes. No experimental data exist for the tobacco protein itself.
Summary: PAINT assertion at PANTHER node PTN001005190, a eukaryote-wide node over plant PRORP1/2/3 and animal and trypanosome PRORPs, seeded by Arabidopsis PRORP1/2/3, Drosophila mldr, human PRORP and two Trypanosoma brucei proteins. The tobacco protein lies inside this clade (PTHR13547:SF13, the same subfamily as Arabidopsis PRORP2) and retains the complete NYN catalytic site.
Reason: Ribonuclease P activity is the defining activity of the family and has been measured directly for the subfamily member Arabidopsis PRORP2, which cleaves nuclear pre-tRNAs in a Mg2+-dependent manner. The target sits within the PAINT clade and has all catalytic aspartates (D310, D386, D387, D405) and zinc ligands of PRORP1 conserved, so there is no indication of loss of activity. Plant PRORPs are active as single proteins, so the activity is properly attributed to the polypeptide.
Our biochemical studies indicate that PRORP2 requires Mg(2+) for catalysis and catalyzes the maturation of nuclear encoded substrates up to 10-fold faster than mitochondrial encoded precursor nad6 t-element under single-turnover conditions.
protein-only RNase P enzymes called PRORP (for proteinaceous RNase P) support RNase P activity in vivo in both organelles and the nucleus in Arabidopsis.
Summary: PAINT assertion at PTN001005190 seeded by Arabidopsis PRORP1, PRORP2 and PRORP3, whose in vivo roles in organellar and nuclear pre-tRNA 5' maturation were shown genetically. The enzyme itself catalyses the step, so this is genuine process participation.
Reason: tRNA 5'-leader removal is the process directly executed by RNase P activity. The tobacco protein is in the PRORP2 subfamily and is catalytically intact. The seed set includes the subfamily member PRORP2, for which nuclear RNase P function in vivo is documented.
thaliana PRORP2 and PRORP3 can catalyze the maturation of the 5′ ends of two nuclear pre-tRNA substrates in vivo, confirming their predicted roles as RNase Ps in the nucleus
Summary: Combined automated annotation (ARBA:ARBA00089897 with PANTHER node PTN004425149) for the same process as the IBA row. The two rows make the same claim.
Reason: Same claim as the accepted IBA row, independently reached through the PANTHER family assignment; supported by the Arabidopsis PRORP2 evidence and the conserved catalytic site.
thaliana PRORP2 and PRORP3 can catalyze the maturation of the 5′ ends of two nuclear pre-tRNA substrates in vivo, confirming their predicted roles as RNase Ps in the nucleus
Our biochemical studies indicate that PRORP2 requires Mg(2+) for catalysis and catalyzes the maturation of nuclear encoded substrates up to 10-fold faster than mitochondrial encoded precursor nad6 t-element under single-turnover conditions.
ISS PMID:26655022 Nuclear Protein-Only Ribonuclease P2 Structure and Biochemic...
NEW
Summary: Proposed location. The tobacco protein is in PANTHER subfamily PTHR13547:SF13 with Arabidopsis PRORP2, a nuclear protein, and is about equally close to the other nuclear paralog PRORP3; it is more distant from the organellar PRORP1 (SF1) and has no N-terminal transit-peptide extension.
Reason: Compartment is the property that distinguishes the plant PRORP subfamilies (PRORP1 organellar, PRORP2/PRORP3 nuclear), so subfamily membership plus absence of a targeting presequence supports nuclear localisation. PAINT made no cellular-component assertion for the plant PRORP nodes, so this does not contradict a curator decision. Direct localisation in tobacco has not been tested.
extension corresponding to the PRORP1 organellar transit peptide
Core Functions
Single-protein RNase P of the PRORP2 subfamily that cleaves the 5' leader from precursor tRNAs using a Mg2+-dependent NYN metallonuclease active site, with substrate recognition by the PPR region; predicted to act on nuclear-encoded pre-tRNAs in the nucleus.
Our biochemical studies indicate that PRORP2 requires Mg(2+) for catalysis and catalyzes the maturation of nuclear encoded substrates up to 10-fold faster than mitochondrial encoded precursor nad6 t-element under single-turnover conditions.
Q: Tobacco is allotetraploid; does A0A1S3Y076 have a homeolog, and are both copies of the nuclear PRORP expressed and functionally redundant, as Arabidopsis PRORP2 and PRORP3 are?
Q: Does the tobacco nuclear PRORP associate with TRM1-type m2,2G26 methyltransferases as Arabidopsis PRORP2 does?
Suggested Experiments
Experiment: Fluorescent-protein fusion of A0A1S3Y076 transiently expressed in N. benthamiana or tobacco leaves, imaged with nuclear, plastid and mitochondrial markers.
Hypothesis: A0A1S3Y076 localises to the nucleus and not to organelles, as expected for a PRORP2-subfamily protein lacking a transit peptide.
Experiment: In vitro cleavage assay with recombinant A0A1S3Y076 on a nuclear-encoded pre-tRNA, with a D386A/D387A catalytic mutant as negative control.
Hypothesis: The protein cleaves the 5' leader at the canonical RNase P site in a Mg2+-dependent manner and the catalytic mutant is inactive.
External Prediction Reviews
These computational predictions are reviewed separately from the GOA annotation set used for this review. The assessments below are from this project and do not constitute official GO annotations or endorsement by GO/UniProt. They are not included in the existing annotation review above.
Nuclear-type proteinaceous RNase P (PRORP2 subfamily) of tobacco. Its single ProtNLM2 prediction, tRNA processing, is correct but is an ancestor of the accepted tRNA 5'-leader removal annotation.
LSP — Less precise than existing annotation Review score: 2/2
Prediction method: ProtNLM2 · Version: UniProt 2024_06 pilot
Review rationale: The target is a catalytically intact PRORP in PANTHER subfamily PTHR13547:SF13 with Arabidopsis PRORP2, and the gene already carries tRNA 5'-leader removal (GO:0001682, IBA and IEA), which the gene review accepts. Arabidopsis PRORP2 and PRORP3 mature nuclear pre-tRNA 5' ends in vivo (PMID:26655022) and PRORPs supply RNase P activity in plants (PMID:22549728), justifying the subfamily transfer. GO:0008033 tRNA processing is an is_a ancestor of GO:0001682 (OLS4), so the prediction is correct but less specific than the existing supported annotation. No tobacco-specific experimental literature was found in PubMed.
PMID:26655022: "thaliana PRORP2 and PRORP3 can catalyze the maturation of the 5′ ends of two nuclear pre-tRNA substrates in vivo, confirming their predicted roles as RNase Ps in the nucleus"
PMID:22549728: "cleaves the 5' leader sequence of tRNA precursors"