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PMID:16807317

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Citation

de la Fuente van Bentem, S, Anrather, D, Roitinger, E, Djamei, A, Hufnagl, T, Barta, A, Csaszar, E, Dohnal, I, Lecourieux, D and Hirt, H (2006) Phosphoproteomics reveals extensive in vivo phosphorylation of Arabidopsis proteins involved in RNA metabolism. Nucleic Acids Res. 34:3267-78

Abstract

Most regulatory pathways are governed by the reversible phosphorylation of proteins. Recent developments in mass spectrometry-based technology allow the large-scale analysis of protein phosphorylation. Here, we show the application of immobilized metal affinity chromatography to purify phosphopeptides from Arabidopsis extracts. Phosphopeptide sequences were identified by liquid chromatography-tandem mass spectrometry (LC-MS/MS/MS). A total of 79 unique phosphorylation sites were determined in 22 phosphoproteins with a putative role in RNA metabolism, including splicing of mRNAs. Among these phosphoproteins, 12 Ser/Arg-rich (SR) splicing factors were identified. A conserved phosphorylation site was found in most of the phosphoproteins, including the SR proteins, suggesting that these proteins are targeted by the same or a highly related protein kinase. To test this hypothesis, Arabidopsis SR protein-specific kinase 4 (SRPK4) that was initially identified as an interactor of SR proteins was tested for its ability to phosphorylate the SR protein RSp31. In vitro kinase assays showed that all in vivo phosphorylation sites of RSp31 were targeted by SRPK4. These data suggest that the plant mRNA splicing machinery is a major target of phosphorylation and that a considerable number of proteins involved in RNA metabolism may be targeted by SRPKs.

Links

PubMed PMC1904105 Online version:10.1093/nar/gkl429

Keywords

Amino Acid Motifs; Amino Acid Sequence; Arabidopsis/enzymology; Arabidopsis/metabolism; Arabidopsis Proteins/chemistry; Arabidopsis Proteins/metabolism; Cell Nucleus/metabolism; Chromatography, Affinity; Cytosol/metabolism; Mass Spectrometry; Molecular Sequence Data; Phosphopeptides/chemistry; Phosphopeptides/isolation & purification; Phosphoproteins/chemistry; Phosphoproteins/metabolism; Phosphorylation; Protein Isoforms/metabolism; Protein-Serine-Threonine Kinases/metabolism; Proteomics/methods; RNA Splicing; RNA, Messenger/metabolism; RNA, Plant/metabolism; RNA-Binding Proteins/metabolism; Sequence Homology, Amino Acid

Significance

Annotations

Gene product Qualifier GO Term Evidence Code with/from Aspect Extension Notes Status

ARATH:SR34A

acts_upstream_of_or_within

GO:0008380: RNA splicing

ECO:0000303: author statement without traceable support used in manual assertion

P

Seeded From UniProt

complete

ARATH:F4I448

acts_upstream_of_or_within

GO:0048510: regulation of timing of transition from vegetative to reproductive phase

ECO:0000315: mutant phenotype evidence used in manual assertion

P

Seeded From UniProt

complete

ARATH:F4J982

acts_upstream_of_or_within

GO:0006468: protein phosphorylation

ECO:0000314: direct assay evidence used in manual assertion

P

Seeded From UniProt

complete

ARATH:F4J982

enables

GO:0004672: protein kinase activity

ECO:0000314: direct assay evidence used in manual assertion

F

Seeded From UniProt

complete

ARATH:RSZ22

acts_upstream_of_or_within

GO:0008380: RNA splicing

ECO:0000303: author statement without traceable support used in manual assertion

P

Seeded From UniProt

complete

ARATH:SRS31

acts_upstream_of_or_within

GO:0008380: RNA splicing

ECO:0000303: author statement without traceable support used in manual assertion

P

Seeded From UniProt

complete

ARATH:RS40

acts_upstream_of_or_within

GO:0008380: RNA splicing

ECO:0000303: author statement without traceable support used in manual assertion

P

Seeded From UniProt

complete

ARATH:SRS41

acts_upstream_of_or_within

GO:0008380: RNA splicing

ECO:0000303: author statement without traceable support used in manual assertion

P

Seeded From UniProt

complete

ARATH:SR45A

acts_upstream_of_or_within

GO:0008380: RNA splicing

ECO:0000303: author statement without traceable support used in manual assertion

P

Seeded From UniProt

complete

ARATH:SRC30

acts_upstream_of_or_within

GO:0008380: RNA splicing

ECO:0000303: author statement without traceable support used in manual assertion

P

Seeded From UniProt

complete

ARATH:Q8L7W3

acts_upstream_of_or_within

GO:0008380: RNA splicing

ECO:0000303: author statement without traceable support used in manual assertion

P

Seeded From UniProt

complete

ARATH:ESP3

acts_upstream_of_or_within

GO:0008380: RNA splicing

ECO:0000303: author statement without traceable support used in manual assertion

P

Seeded From UniProt

complete

ARATH:RSZ33

acts_upstream_of_or_within

GO:0008380: RNA splicing

ECO:0000303: author statement without traceable support used in manual assertion

P

Seeded From UniProt

complete

ARATH:SC35

acts_upstream_of_or_within

GO:0008380: RNA splicing

ECO:0000303: author statement without traceable support used in manual assertion

P

Seeded From UniProt

complete

ARATH:RSZ32

acts_upstream_of_or_within

GO:0008380: RNA splicing

ECO:0000303: author statement without traceable support used in manual assertion

P

Seeded From UniProt

complete

ARATH:SL30A

acts_upstream_of_or_within

GO:0008380: RNA splicing

ECO:0000303: author statement without traceable support used in manual assertion

P

Seeded From UniProt

complete

ARATH:SF1

acts_upstream_of_or_within

GO:0008380: RNA splicing

ECO:0000303: author statement without traceable support used in manual assertion

P

Seeded From UniProt

complete

ARATH:CYP63

acts_upstream_of_or_within

GO:0008380: RNA splicing

ECO:0000303: author statement without traceable support used in manual assertion

P

Seeded From UniProt

complete

ARATH:BTR1

acts_upstream_of_or_within

GO:0008380: RNA splicing

ECO:0000303: author statement without traceable support used in manual assertion

P

Seeded From UniProt

complete

ARATH:SR45

acts_upstream_of_or_within

GO:0008380: RNA splicing

ECO:0000303: author statement without traceable support used in manual assertion

P

Seeded From UniProt

complete

ARATH:SRC33

acts_upstream_of_or_within

GO:0008380: RNA splicing

ECO:0000303: author statement without traceable support used in manual assertion

P

Seeded From UniProt

complete

ARATH:RZ22A

acts_upstream_of_or_within

GO:0008380: RNA splicing

ECO:0000303: author statement without traceable support used in manual assertion

P

Seeded From UniProt

complete

ARATH:Q9LU44

involved_in

GO:0008380: RNA splicing

ECO:0000303: author statement without traceable support used in manual assertion

P

Seeded From UniProt

complete

See also

References

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