title: Phosphoproteomic analysis identifies the tumor suppressor PDCD4 as a RSK substrate negatively regulated by 14-3-3.
abstract: The Ras/MAPK signaling cascade regulates various biological functions, including cell growth and proliferation. As such, this pathway is frequently deregulated in several types of cancer, including most cases of melanoma. RSK (p90 ribosomal S6 kinase) is a MAPK-activated protein kinase required for melanoma growth and proliferation, but relatively little is known about its exact function and the nature of its substrates. Herein, we used a quantitative phosphoproteomics approach to define the signaling networks regulated by RSK in melanoma. To more accurately predict direct phosphorylation substrates, we defined the RSK consensus phosphorylation motif and found significant overlap with the binding consensus of 14-3-3 proteins. We thus characterized the phospho-dependent 14-3-3 interactome in melanoma cells and found that a large proportion of 14-3-3 binding proteins are also potential RSK substrates. Our results show that RSK phosphorylates the tumor suppressor PDCD4 (programmed cell death protein 4) on two serine residues (Ser76 and Ser457) that regulate its subcellular localization and interaction with 14-3-3 proteins. We found that 14-3-3 binding promotes PDCD4 degradation, suggesting an important role for RSK in the inactivation of PDCD4 in melanoma. In addition to this tumor suppressor, our results suggest the involvement of RSK in a vast array of unexplored biological functions with relevance in oncogenesis.
authors.affiliation: Institute for Research in Immunology and Cancer, Montreal, QC, Canada H3C 3J7;Department of Pathology and Cell Biology, Faculty of Medicine, Université de Montréal, Montreal, QC, Canada H3C 3J7;
authors.forename: Jacob A
authors.initials: JA
authors.lastname: Galan
authors.affiliation: Department of Cell Biology, Harvard Medical School, Boston, MA 02115;
authors.forename: Kathryn M
authors.initials: KM
authors.lastname: Geraghty
authors.affiliation: Institute for Research in Immunology and Cancer, Montreal, QC, Canada H3C 3J7;
authors.forename: Geneviève
authors.initials: G
authors.lastname: Lavoie
authors.affiliation: Institute for Research in Immunology and Cancer, Montreal, QC, Canada H3C 3J7;Department of Chemistry, Faculty of Arts and Science, Université de Montréal, Montreal, QC, Canada H3C 3J7;
authors.forename: Evgeny
authors.initials: E
authors.lastname: Kanshin
authors.affiliation: Institute for Research in Immunology and Cancer, Montreal, QC, Canada H3C 3J7;Department of Pathology and Cell Biology, Faculty of Medicine, Université de Montréal, Montreal, QC, Canada H3C 3J7;
authors.forename: Joseph
authors.initials: J
authors.lastname: Tcherkezian
authors.affiliation: Institute for Research in Immunology and Cancer, Montreal, QC, Canada H3C 3J7;Department of Pathology and Cell Biology, Faculty of Medicine, Université de Montréal, Montreal, QC, Canada H3C 3J7;
authors.forename: Viviane
authors.initials: V
authors.lastname: Calabrese
authors.affiliation: Department of Pharmacology, Yale University School of Medicine, New Haven, CT 06510; and.
authors.forename: Grace R
authors.initials: GR
authors.lastname: Jeschke
authors.affiliation: Department of Pharmacology, Yale University School of Medicine, New Haven, CT 06510; and.
authors.forename: Benjamin E
authors.initials: BE
authors.lastname: Turk
authors.affiliation: Department of Biology, University of Vermont, Burlington, VT 05405.
authors.forename: Bryan A
authors.initials: BA
authors.lastname: Ballif
authors.affiliation: Department of Cell Biology, Harvard Medical School, Boston, MA 02115;
authors.forename: John
authors.initials: J
authors.lastname: Blenis
authors.affiliation: Institute for Research in Immunology and Cancer, Montreal, QC, Canada H3C 3J7;Department of Chemistry, Faculty of Arts and Science, Université de Montréal, Montreal, QC, Canada H3C 3J7;
authors.forename: Pierre
authors.initials: P
authors.lastname: Thibault
authors.affiliation: Institute for Research in Immunology and Cancer, Montreal, QC, Canada H3C 3J7;Department of Pathology and Cell Biology, Faculty of Medicine, Université de Montréal, Montreal, QC, Canada H3C 3J7; philippe.roux@umontreal.ca.
authors.forename: Philippe P
authors.initials: PP
authors.lastname: Roux
mesh_terms: D048948:14-3-3 Proteins / Q000378:metabolism; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism*; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D016384:Consensus Sequence; D006801:Humans; D008545:Melanoma / Q000378:metabolism / Q000473:pathology; D008954:Models, Biological; D008969:Molecular Sequence Data; D019151:Peptide Library; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D010768:Phosphoserine / Q000378:metabolism; D011485:Protein Binding; D021381:Protein Transport; D059748:Proteolysis; D020543:Proteome / Q000378:metabolism; D040901:Proteomics / Q000379:methods*; D016601:RNA-Binding Proteins / Q000378:metabolism*; D038744:Ribosomal Protein S6 Kinases, 90-kDa / Q000378:metabolism*; D013379:Substrate Specificity; D025521:Tumor Suppressor Proteins / Q000378:metabolism*
chemical_list: D048948:14-3-3 Proteins; D051017:Apoptosis Regulatory Proteins; C114487:PDCD4 protein, human; D019151:Peptide Library; D010750:Phosphoproteins; D020543:Proteome; D016601:RNA-Binding Proteins; D025521:Tumor Suppressor Proteins; D010768:Phosphoserine; D038744:Ribosomal Protein S6 Kinases, 90-kDa