title: Opposing effects of pericentrin and microcephalin on the pericentriolar material regulate CHK1 activation in the DNA damage response.
abstract: Genotoxic stresses lead to centrosome amplification, a frequently-observed feature in cancer that may contribute to genome instability and to tumour cell invasion. Here we have explored how the centrosome controls DNA damage responses. For most of the cell cycle, centrosomes consist of two centrioles embedded in the proteinaceous pericentriolar material (PCM). Recent data indicate that the PCM is not an amorphous assembly of proteins, but actually a highly organised scaffold around the centrioles. The large coiled-coil protein, pericentrin, participates in PCM assembly and has been implicated in the control of DNA damage responses (DDRs) through its interactions with checkpoint kinase 1 (CHK1) and microcephalin (MCPH1). CHK1 is required for DNA damage-induced centrosome amplification, whereas MCPH1 deficiency greatly increases the amplification seen after DNA damage. We found that the PCM showed a marked expansion in volume and a noticeable change in higher-order organisation after ionising radiation treatment. PCM expansion was dependent on CHK1 kinase activity and was potentiated by MCPH1 deficiency. Furthermore, pericentrin deficiency or mutation of a separase cleavage site blocked DNA damage-induced PCM expansion. The extent of nuclear CHK1 activation after DNA damage reflected the level of PCM expansion, with a reduction in pericentrin-deficient or separase cleavage site mutant-expressing cells, and an increase in MCPH1-deficient cells that was suppressed by the loss of pericentrin. Deletion of the nuclear export signal of CHK1 led to its hyperphosphorylation after irradiation and reduced centrosome amplification. Deletion of the nuclear localisation signal led to low CHK1 activation and low centrosome amplification. From these data, we propose a feedback loop from the PCM to the nuclear DDR in which CHK1 regulates pericentrin-dependent PCM expansion to control its own activation.
authors.affiliation: Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, Galway, Ireland.
authors.forename: A K
authors.initials: AK
authors.lastname: Antonczak
authors.affiliation: Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, Galway, Ireland.
authors.forename: L I
authors.initials: LI
authors.lastname: Mullee
authors.affiliation: Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, Galway, Ireland.
authors.forename: Y
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Lunenfeld-Tanenbaum Research Institute, University of Toronto, Toronto, Canada.
authors.forename: D
authors.initials: D
authors.lastname: Comartin
authors.affiliation: Department of Cell Biology, Johns Hopkins University, Baltimore, MD, USA.
authors.forename: T
authors.initials: T
authors.lastname: Inoue
authors.affiliation: Lunenfeld-Tanenbaum Research Institute, University of Toronto, Toronto, Canada.
authors.forename: L
authors.initials: L
authors.lastname: Pelletier
authors.affiliation: Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, Galway, Ireland.
authors.forename: C G
authors.initials: CG
authors.lastname: Morrison
mesh_terms: D000818:Animals; D000941:Antigens / Q000235:genetics / Q000502:physiology*; D001665:Binding Sites; D018797:Cell Cycle Proteins / Q000235:genetics / Q000502:physiology*; D045744:Cell Line, Tumor; D002502:Centrioles / Q000378:metabolism / Q000648:ultrastructure; D018385:Centrosome / Q000502:physiology* / Q000528:radiation effects / Q000648:ultrastructure; D000071877:Checkpoint Kinase 1; D002645:Chickens; D004249:DNA Damage; D004789:Enzyme Activation; D025461:Feedback, Physiological; D017930:Genes, Reporter; D009154:Mutation; D009363:Neoplasm Proteins / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000172:deficiency / Q000235:genetics / Q000502:physiology*; D034521:Nuclear Matrix-Associated Proteins / Q000235:genetics / Q000502:physiology*; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D011499:Protein Processing, Post-Translational / Q000502:physiology*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D064247:Separase / Q000378:metabolism
chemical_list: D000941:Antigens; D018797:Cell Cycle Proteins; D009363:Neoplasm Proteins; D009419:Nerve Tissue Proteins; D034521:Nuclear Matrix-Associated Proteins; D011993:Recombinant Fusion Proteins; C086178:pericentrin; D011494:Protein Kinases; D000071877:Checkpoint Kinase 1; D064247:Separase