Identification of Targets and Interaction Partners of Arginyl-tRNA Protein Transferase in the Moss Physcomitrella patens.

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The lack of arginylation causes severe developmental defects in moss, affects the low oxygen response in Arabidopsis thaliana and is embryo lethal in Drosophila and in mice. Although several studies investigated impact and function of the responsible enzyme, the arginyl-tRNA protein transferase (ATE) in plants, identification of arginylated proteins by mass spectrometry was not hitherto achieved. In the present study, we report the identification of targets and interaction partners of ATE in the model plant Physcomitrella patens by mass spectrometry, employing two different immuno-affinity strategies and a recently established transgenic ATE:GUS reporter line (Schuessele et al., 2016 New Phytol. , DOI: 10.1111/nph.13656). Here we use a commercially available antibody against the fused reporter protein (β-glucuronidase) to pull down ATE and its interacting proteins and validate its in vivo interaction with a class I small heatshock protein via Förster resonance energy transfer (FRET). Additionally, we apply and modify a method that already successfully identified arginylated proteins from mouse proteomes by using custom-made antibodies specific for N-terminal arginine. As a result, we identify four arginylated proteins from Physcomitrella patens with high confidence.Data are available via ProteomeXchange with identifier PXD003228 and PXD003232.", "claims": [{"id": "claim-target-result", "statement": "The paper identifies four arginylated proteins from Physcomitrella patens with high confidence, but it does not state a statistical method for this registered target; the source data are available through ProteomeXchange identifiers PXD003228 and PXD003232.", "kind": "qualitative", "target": null, "evidence": [{"asset": "source:pmid:27067052", "locator": "sections[0] \"Abstract\" paragraph 1", "quote": "As a result, we identify four arginylated proteins from Physcomitrella patens with high confidence."}, {"asset": "source:pmid:27067052", "locator": "sections[0] \"Abstract\" paragraph 2", "quote": "Data are available via ProteomeXchange with identifier PXD003228 and PXD003232."}], "state": "complete"}], "gaps": []}, "experiment": {"state": "partial", "objective": "The paper identifies four arginylated proteins from Physcomitrella patens with high confidence, but it does not state a statistical method for this registered target; the source data are available through ProteomeXchange identifiers PXD003228 and PXD003232.", "steps": [{"id": "read_published_estimate", "order": 1, "name": "Read the published estimate from the source", "kind": "other", "state": "complete", "blocked_on": null, "description": "The claim, method and evidence span published with the source article.", "duration_hours": null, "parameters": {}, "inputs": ["source:pmid:27067052"], "outputs": ["the published estimate and its evidence"], "instruments": [], "reagents": []}, {"id": "obtain_source_data", "order": 2, "name": "Obtain the source data from the named repository", "kind": "data-preparation", "state": "blocked", "blocked_on": "data:accession_absent", "description": "The GEO series accession resolves and its sample accessions are listed. 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