title: Cellular abundance of Mps1 and the role of its carboxyl terminal tail in substrate recruitment.
abstract: Mps1 is a protein kinase that regulates normal mitotic progression and the spindle checkpoint in response to spindle damage. The levels of Mps1 are relatively low in cells during interphase but elevated in mitosis or upon activation of the spindle checkpoint, although the dynamic range of Mps1 expression and the Mps1 catalytic mechanism have not been carefully characterized. Our recent structural studies of the Mps1 kinase domain revealed that the carboxyl-terminal tail region of Mps1 is unstructured, raising the question of whether this region has any functional role in Mps1 catalysis. Here we first determined the cellular abundance of Mps1 during cell cycle progression and found that Mps1 levels vary between 60,000 per cell in early G(1) and 110,000 per cell during mitosis. We studied phosphorylation of a number of Mps1 substrates in vitro and in culture cells. Unexpectedly, we found that the unstructured carboxyl-terminal region of Mps1 plays an essential role in substrate recruitment. Kinetics studies using the purified recombinant wild type and mutant kinases indicate that the carboxyl-terminal tail is largely dispensable for autophosphorylation of Mps1 but critical for trans-phosphorylation of substrates in vitro and in cultured cells. Mps1 mutant without the unstructured tail region is defective in mediating spindle assembly checkpoint activation. Our results underscore the importance of the unstructured tail region of Mps1 in kinase activation.
authors.affiliation: Department of Chemistry and Biochemistry, University of Colorado, Boulder, Colorado 80309, USA.
authors.forename: Tingting
authors.initials: T
authors.lastname: Sun
authors.forename: Xiaomei
authors.initials: X
authors.lastname: Yang
authors.forename: Wei
authors.initials: W
authors.lastname: Wang
authors.forename: Xiaojuan
authors.initials: X
authors.lastname: Zhang
authors.forename: Quanbin
authors.initials: Q
authors.lastname: Xu
authors.forename: Songcheng
authors.initials: S
authors.lastname: Zhu
authors.forename: Robert
authors.initials: R
authors.lastname: Kuchta
authors.forename: Guanjun
authors.initials: G
authors.lastname: Chen
authors.forename: Xuedong
authors.initials: X
authors.lastname: Liu
mesh_terms: D002384:Catalysis; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D004789:Enzyme Activation / Q000502:physiology; D016193:G1 Phase / Q000502:physiology*; D006367:HeLa Cells; D006801:Humans; D008938:Mitosis / Q000502:physiology*; D009154:Mutation; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D011505:Protein-Tyrosine Kinases; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D008941:Spindle Apparatus / Q000235:genetics / Q000378:metabolism*
chemical_list: D018797:Cell Cycle Proteins; D011994:Recombinant Proteins; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; C075840:TTK protein, human