title: Identification of the PLK2-dependent phosphopeptidome by quantitative proteomics [corrected].
abstract: Polo-like kinase 2 (PLK2) has been recently recognized as the major enzyme responsible for phosphorylation of α-synuclein at S129 in vitro and in vivo, suggesting that this kinase may play a key role in the pathogenesis of Parkinson's disease and other synucleinopathies. Moreover PLK2 seems to be implicated in cell division, oncogenesis, and synaptic regulation of the brain. However little is known about the phosphoproteome generated by PLK2 and, consequently the overall impact of PLK2 on cellular signaling. To fill this gap we exploited an approach based on in vitro kinase assay and quantitative phosphoproteomics. A proteome-derived peptide library obtained by digestion of undifferentiated human neuroblastoma cell line was exhaustively dephosphorylated by lambda phosphatase followed by incubation with or without PLK2 recombinant kinase. Stable isotope labeling based quantitative phosphoproteomics was applied to identify the phosphosites generated by PLK2. A total of 98 unique PLK2-dependent phosphosites from 89 proteins were identified by LC-MS/MS. Analysis of the primary structure of the identified phosphosites allowed the detailed definition of the kinase specificity and the compilation of a list of potential PLK2 targets among those retrieved in PhosphositePlus, a curated database of in cell/vivo phosphorylation sites.
authors.affiliation: Department of Biomedical Sciences, University of Padova, Padova, Italy; Proteomics Center of Padova University, Padova, Italy.
authors.forename: Cinzia
authors.initials: C
authors.lastname: Franchin
authors.affiliation: Department of Biomedical Sciences, University of Padova, Padova, Italy.
authors.forename: Luca
authors.initials: L
authors.lastname: Cesaro
authors.affiliation: Department of Biomedical Sciences, University of Padova, Padova, Italy; CNR Institute of Neurosciences, Padova, Italy.
authors.forename: Lorenzo A
authors.initials: LA
authors.lastname: Pinna
authors.affiliation: Department of Biomedical Sciences, University of Padova, Padova, Italy; Proteomics Center of Padova University, Padova, Italy.
authors.forename: Giorgio
authors.initials: G
authors.lastname: Arrigoni
authors.affiliation: Department of Biomedical Sciences, University of Padova, Padova, Italy.
authors.forename: Mauro
authors.initials: M
authors.lastname: Salvi
mesh_terms: D000595:Amino Acid Sequence; D020134:Catalytic Domain; D045744:Cell Line, Tumor; D016384:Consensus Sequence; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010750:Phosphoproteins / Q000737:chemistry / Q000378:metabolism; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D017346:Protein Serine-Threonine Kinases / Q000737:chemistry / Q000502:physiology*; D020543:Proteome / Q000737:chemistry / Q000378:metabolism*; D053719:Tandem Mass Spectrometry
chemical_list: D010750:Phosphoproteins; D020543:Proteome; C496126:PLK2 protein, human; D017346:Protein Serine-Threonine Kinases