title: Spatial exclusivity combined with positive and negative selection of phosphorylation motifs is the basis for context-dependent mitotic signaling.
abstract: The timing and localization of events during mitosis are controlled by the regulated phosphorylation of proteins by the mitotic kinases, which include Aurora A, Aurora B, Nek2 (never in mitosis kinase 2), Plk1 (Polo-like kinase 1), and the cyclin-dependent kinase complex Cdk1/cyclin B. Although mitotic kinases can have overlapping subcellular localizations, each kinase appears to phosphorylate its substrates on distinct sites. To gain insight into the relative importance of local sequence context in kinase selectivity, identify previously unknown substrates of these five mitotic kinases, and explore potential mechanisms for substrate discrimination, we determined the optimal substrate motifs of these major mitotic kinases by positional scanning oriented peptide library screening (PS-OPLS). We verified individual motifs with in vitro peptide kinetic studies and used structural modeling to rationalize the kinase-specific selection of key motif-determining residues at the molecular level. Cross comparisons among the phosphorylation site selectivity motifs of these kinases revealed an evolutionarily conserved mutual exclusion mechanism in which the positively and negatively selected portions of the phosphorylation motifs of mitotic kinases, together with their subcellular localizations, result in proper substrate targeting in a coordinated manner during mitosis.
authors.affiliation: Koch Institute for Integrative Cancer Research, Department of Biology, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.
authors.forename: Jes
authors.initials: J
authors.lastname: Alexander
authors.forename: Daniel
authors.initials: D
authors.lastname: Lim
authors.forename: Brian A
authors.initials: BA
authors.lastname: Joughin
authors.forename: Björn
authors.initials: B
authors.lastname: Hegemann
authors.forename: James R A
authors.initials: JR
authors.lastname: Hutchins
authors.forename: Tobias
authors.initials: T
authors.lastname: Ehrenberger
authors.forename: Frank
authors.initials: F
authors.lastname: Ivins
authors.forename: Fabio
authors.initials: F
authors.lastname: Sessa
authors.forename: Otto
authors.initials: O
authors.lastname: Hudecz
authors.forename: Erich A
authors.initials: EA
authors.lastname: Nigg
authors.forename: Andrew M
authors.initials: AM
authors.lastname: Fry
authors.forename: Andrea
authors.initials: A
authors.lastname: Musacchio
authors.forename: P Todd
authors.initials: PT
authors.lastname: Stukenberg
authors.forename: Karl
authors.initials: K
authors.lastname: Mechtler
authors.forename: Jan-Michael
authors.initials: JM
authors.lastname: Peters
authors.forename: Stephen J
authors.initials: SJ
authors.lastname: Smerdon
authors.forename: Michael B
authors.initials: MB
authors.lastname: Yaffe
mesh_terms: D020816:Amino Acid Motifs; D000818:Animals; D019143:Evolution, Molecular; D006801:Humans; D008938:Mitosis / Q000502:physiology*; D019151:Peptide Library; D010766:Phosphorylation / Q000502:physiology; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*; D029867:Xenopus Proteins / Q000378:metabolism*; D014982:Xenopus laevis
chemical_list: D019151:Peptide Library; D029867:Xenopus Proteins; D017346:Protein Serine-Threonine Kinases