title: Systematic phosphorylation analysis of human mitotic protein complexes.
abstract: Progression through mitosis depends on a large number of protein complexes that regulate the major structural and physiological changes necessary for faithful chromosome segregation. Most, if not all, of the mitotic processes are regulated by a set of mitotic protein kinases that control protein activity by phosphorylation. Although many mitotic phosphorylation events have been identified in proteome-scale mass spectrometry studies, information on how these phosphorylation sites are distributed within mitotic protein complexes and which kinases generate these phosphorylation sites is largely lacking. We used systematic protein-affinity purification combined with mass spectrometry to identify 1818 phosphorylation sites in more than 100 mitotic protein complexes. In many complexes, the phosphorylation sites were concentrated on a few subunits, suggesting that these subunits serve as "switchboards" to relay the kinase-regulatory signals within the complexes. Consequent bioinformatic analyses identified potential kinase-substrate relationships for most of these sites. In a subsequent in-depth analysis of key mitotic regulatory complexes with the Aurora kinase B (AURKB) inhibitor Hesperadin and a new Polo-like kinase (PLK1) inhibitor, BI 4834, we determined the kinase dependency for 172 phosphorylation sites on 41 proteins. Combination of the results of the cellular studies with Scansite motif prediction enabled us to identify 14 sites on six proteins as direct candidate substrates of AURKB or PLK1.
authors: Research Institute of Molecular Pathology, Dr. Bohr-Gasse 7, 1030 Vienna, Austria.
authors: Björn
authors: B
authors: Hegemann
authors: James R A
authors: JR
authors: Hutchins
authors: Otto
authors: O
authors: Hudecz
authors: Maria
authors: M
authors: Novatchkova
authors: Jonathan
authors: J
authors: Rameseder
authors: Martina M
authors: MM
authors: Sykora
authors: Sihan
authors: S
authors: Liu
authors: Michael
authors: M
authors: Mazanek
authors: Péter
authors: P
authors: Lénárt
authors: Jean-Karim
authors: JK
authors: Hériché
authors: Ina
authors: I
authors: Poser
authors: Norbert
authors: N
authors: Kraut
authors: Anthony A
authors: AA
authors: Hyman
authors: Michael B
authors: MB
authors: Yaffe
authors: Karl
authors: K
authors: Mechtler
authors: Jan-Michael
authors: JM
authors: Peters
mesh_terms: D018797:Cell Cycle Proteins / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D008938:Mitosis / Q000502:physiology*; D010766:Phosphorylation / Q000502:physiology; D011494:Protein Kinases / Q000378:metabolism*
chemical_list: D018797:Cell Cycle Proteins; D011494:Protein Kinases