leupeptins and hesperadin

Error-free chromosome segregation depends on the precise regulation of phosphorylation to stabilize kinetochore-microtubule attachments (K-fibres) on sister chromatids that have attached to opposite spindle poles (bi-oriented). In many instances, phosphorylation correlates with K-fibre destabilization. Consistent with this, multiple kinases, including Aurora B and Plk1, are enriched at kinetochores of mal-oriented chromosomes when compared with bi-oriented chromosomes, which have stable attachments. Paradoxically, however, these kinases also target to prometaphase chromosomes that have not yet established spindle attachments and it is therefore unclear how kinetochore-microtubule interactions can be stabilized when kinase levels are high. Here we show that the generation of stable K-fibres depends on the B56-PP2A phosphatase, which is enriched at centromeres/kinetochores of unattached chromosomes. When B56-PP2A is depleted, K-fibres are destabilized and chromosomes fail to align at the spindle equator. Strikingly, B56-PP2A depletion increases the level of phosphorylation of Aurora B and Plk1 kinetochore substrates as well as Plk1 recruitment to kinetochores. Consistent with increased substrate phosphorylation, we find that chemical inhibition of Aurora or Plk1 restores K-fibres in B56-PP2A-depleted cells. Our findings reveal that PP2A, an essential tumour suppressor, tunes the balance of phosphorylation to promote chromosome-spindle interactions during cell division.

Topics: Aurora Kinase B; Aurora Kinases; Benzamides; Cell Cycle Proteins; Chromosome Segregation; HeLa Cells; Humans; Indoles; Kinetochores; Leupeptins; Microscopy, Fluorescence; Microtubules; Nocodazole; Phosphorylation; Polo-Like Kinase 1; Protein Kinase Inhibitors; Protein Phosphatase 2; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Pteridines; Quinazolines; Recombinant Fusion Proteins; RNA Interference; Sulfonamides; Time Factors; Transfection; Tumor Suppressor Proteins