Proceedings of the National Academy of Sciences - PNAS, 2015-07, Vol.112 (27), p.8481-8486
2015
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- Autor(en) / Beteiligte
- Titel
- Phosphorylation is an on/off switch for 5-hydroxyconiferaldehyde O-methyltransferase activity in poplar monolignol biosynthesis
- Ist Teil von
- Proceedings of the National Academy of Sciences - PNAS, 2015-07, Vol.112 (27), p.8481-8486
- Ort / Verlag
- United States: National Academy of Sciences
- Erscheinungsjahr
- 2015
- Quelle
- MEDLINE
- Beschreibungen/Notizen
- Although phosphorylation has long been known to be an important regulatory modification of proteins, no unequivocal evidence has been presented to show functional control by phosphorylation for the plant monolignol biosynthetic pathway. Here, we present the discovery of phosphorylation-mediated on/off regulation of enzyme activity for 5-hydroxyconiferaldehyde O -methyltransferase 2 (PtrAldOMT2), an enzyme central to monolignol biosynthesis for lignification in stem-differentiating xylem (SDX) of Populus trichocarpa . Phosphorylation turned off the PtrAldOMT2 activity, as demonstrated in vitro by using purified phosphorylated and unphosphorylated recombinant PtrAldOMT2. Protein extracts of P. trichocarpa SDX, which contains endogenous kinases, also phosphorylated recombinant PtrAldOMT2 and turned off the recombinant protein activity. Similarly, ATP/Mn ²âº-activated phosphorylation of SDX protein extracts reduced the endogenous SDX PtrAldOMT2 activity by â¼60%, and dephosphorylation fully restored the activity. Global shotgun proteomic analysis of phosphopeptide-enriched P. trichocarpa SDX protein fractions identified PtrAldOMT2 monophosphorylation at Ser ¹²³ or Ser ¹²ⵠin vivo. Phosphorylation-site mutagenesis verified the PtrAldOMT2 phosphorylation at Ser ¹²³ or Ser ¹²ⵠand confirmed the functional importance of these phosphorylation sites for O -methyltransferase activity. The PtrAldOMT2 Ser ¹²³ phosphorylation site is conserved across 93% of AldOMTs from 46 diverse plant species, and 98% of the AldOMTs have either Ser ¹²³ or Ser ¹²âµ. PtrAldOMT2 is a homodimeric cytosolic enzyme expressed more abundantly in syringyl lignin-rich fiber cells than in guaiacyl lignin-rich vessel cells. The reversible phosphorylation of PtrAldOMT2 is likely to have an important role in regulating syringyl monolignol biosynthesis of P. trichocarpa .
- Sprache
- Englisch
- Identifikatoren
- ISSN: 0027-8424eISSN: 1091-6490DOI: 10.1073/pnas.1510473112Titel-ID: cdi_crossref_primary_10_1073_pnas_1510473112
- Format
- –
- Schlagworte
- Acrolein - analogs & derivatives, Acrolein - metabolism, adenosine triphosphate, AldOMT, Amino Acid Sequence, Binding Sites - genetics, Biocatalysis, biochemical pathways, Biological Sciences, Biosynthesis, Catechols - metabolism, Cells, Chromatography, Liquid, COMT, dephosphorylation, Electrophoresis, Polyacrylamide Gel, enzyme activity, fiber cells, Flowers & plants, Green Fluorescent Proteins - genetics, Green Fluorescent Proteins - metabolism, Isoenzymes - genetics, Isoenzymes - metabolism, lignification, lignin, Lignin - biosynthesis, manganese, Methyltransferases - genetics, Methyltransferases - metabolism, Microscopy, Confocal, Molecular Sequence Data, Mutagenesis, Mutation, Peptides, Phosphoproteins - metabolism, phosphoproteomics, phosphoregulation, Phosphorylation, phosphotransferases (kinases), Plant Proteins - genetics, Plant Proteins - metabolism, Populus - enzymology, Populus - genetics, Populus - metabolism, Populus balsamifera subsp. trichocarpa, Proteins, proteomics, Proteomics - methods, Protoplasts - enzymology, Protoplasts - metabolism, recombinant proteins, Sequence Homology, Amino Acid, Tandem Mass Spectrometry, xylem