Fuller, B. G. et al. Midzone activation of Aurora B in anaphase produces an intracellular phosphorylation gradient. Nature 453, 1132-1136

Department of Biochemistry, University of Virginia School of Medicine, Charlottesville, Virginia 22908, USA.
Nature (Impact Factor: 41.46). 07/2008; 453(7198):1132-6. DOI: 10.1038/nature06923
Source: PubMed


Proper partitioning of the contents of a cell between two daughters requires integration of spatial and temporal cues. The anaphase array of microtubules that self-organize at the spindle midzone contributes to positioning the cell-division plane midway between the segregating chromosomes. How this signalling occurs over length scales of micrometres, from the midzone to the cell cortex, is not known. Here we examine the anaphase dynamics of protein phosphorylation by aurora B kinase, a key mitotic regulator, using fluorescence resonance energy transfer (FRET)-based sensors in living HeLa cells and immunofluorescence of native aurora B substrates. Quantitative analysis of phosphorylation dynamics, using chromosome- and centromere-targeted sensors, reveals that changes are due primarily to position along the division axis rather than time. These dynamics result in the formation of a spatial phosphorylation gradient early in anaphase that is centred at the spindle midzone. This gradient depends on aurora B targeting to a subpopulation of microtubules that activate it. Aurora kinase activity organizes the targeted microtubules to generate a structure-based feedback loop. We propose that feedback between aurora B kinase activation and midzone microtubules generates a gradient of post-translational marks that provides spatial information for events in anaphase and cytokinesis.

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    • "Consistent with this model, photobleaching showed that the CPC binds dynamically to chromosomes in metaphase, but is more static on microtubule bundles during anaphase and cytokinesis (Murata-Hori and Wang, 2002; Delacour-Larose et al., 2004). The CPC generates kinase activity gradients in anaphase, which may promote correct localization (Fuller et al., 2008). Microtubule binding activates the kinase activity of CPC, probably by promoting trans-phosphorylation of the AurkB and INCENP subunits (Kelley et al 2007). "
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    ABSTRACT: Mitotic spindles specify cleavage planes in early embryos by communicating their position and orientation to the cell cortex using microtubule asters that grow out from the spindle poles during anaphase. Chromatin also plays a poorly understood role. Polyspermic fertilization provides a natural experiment where aster pairs from the same spindle (sister asters) have chromatin between them, while asters pairs from different spindles (non-sisters) do not. In frogs, only sister aster pairs induce furrows. We found that only sister asters recruited two conserved furrow-inducing signaling complexes, Chromosome Passenger Complex (CPC) and Centralspindlin, to a plane between them. This explains why only sister pairs induce furrows. We then investigated factors that influenced CPC recruitment to microtubule bundles in intact eggs and a cytokinesis extract system. We found that microtubule stabilization, optimal starting distance between asters, and proximity to chromatin all favored CPC recruitment. We propose a model in which proximity to chromatin biases initial CPC recruitment to microtubules bundles between asters from the same spindle. Next, a positive feedback between CPC recruitment and microtubule stabilization promotes lateral growth of a plane CPC-positive microtubule bundles out to the cortex to position the furrow. © 2015 by The American Society for Cell Biology.
    Molecular biology of the cell 08/2015; DOI:10.1091/mbc.E15-04-0233 · 4.47 Impact Factor
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    • "Protein kinase Donor/acceptor fluorescent Phosphorylation site Phosphorylation binding domain Refs. proteins Cyclin B1-Cdk1 mcerulean/YPet Ser 126 of HsCyclin B1 Polo-box domain of HsPlk1 [19] Aurora-B CyPet/YPet or mTFP1/YPet Thr 67 of HsKif2 FHA2 of ScRad53 [56] [61] Plk1 CyPet/YPet Ser 426 of HsMyt1 FHA2 of ScRad53 [56] Plk1 CyPet/YPet or mTFP1/YPet Ser 17 of c-jun FHA2 of ScRad53 [59] Protein Name of the biosensor Donor/acceptor fluorescent Other domains Refs. proteins ran Rango Cerulean/EYFP Importin-β binding domain [40] of Hs snurportin1 Stathmin/Op18 COPY ECFP/citrine Full length Stathmin/Op18 [54] RhoA Raichu-RhoA SEYFP/SECFP Truncated RhoA and RhoA [73] "
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    ABSTRACT: Mitosis has been studied since the early 1880s as a key event of the cell division cycle where remarkable changes in cellular architecture take place and ultimately lead to an equal segregation of duplicated chromosomes into two daughter cells. A detailed description of the complex and highly ordered cellular events taking place is now available. Many regulators involved in key steps including entry into mitosis, nuclear envelope breakdown, microtubule (MT) spindle formation, and chromosome attachment, as well as mitotic exit and cytokinesis, have also been identified. However, understanding the precise spatio-temporal contribution of each regulator in the cell reorganization process has been technically challenging. This review will focus on a number of recent advances in our understanding of the spatial distribution of protein activities and the temporal regulation of their activation and inactivation during entry and progression through mitosis by the use of intramolecular Förster resonance energy transfer (FRET)-based biosensors.
    Biotechnology Journal 02/2014; 9(2). DOI:10.1002/biot.201300194 · 3.49 Impact Factor
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    • "Aurora B is found at the spindle midzone and at the equatorial cortex during the meta-to-anaphase transition [4]. At the spindle midzone, Aurora B is thought to generate an anaphase phosphorylation gradient toward the cell cortex, which provides spatial information to position the cleavage furrow [5]. In contrast, the importance of cortically localized Aurora B for cytokinesis has remained elusive. "
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    ABSTRACT: Although Aurora B is important in cleavage furrow ingression and completion during cytokinesis, the mechanism by which kinase activity is targeted to the cleavage furrow and the molecule(s) responsible for this process have remained elusive. Here, we demonstrate that an essential mitotic kinesin MKlp2 requires myosin-II for its localization to the equatorial cortex, and this event is required to recruit Aurora B to the equatorial cortex in mammalian cells. This recruitment event is also required to promote the highly focused accumulation of active RhoA at the equatorial cortex and stable ingression of the cleavage furrow in bipolar cytokinesis. Specifically, in drug-induced monopolar cytokinesis, targeting Aurora B to the cell cortex by MKlp2 is essential for cell polarization and furrow formation. Once the furrow has formed, MKlp2 further recruits Aurora B to the growing furrow. This process together with continuous Aurora B kinase activity at the growing furrow is essential for stable furrow propagation and completion. In contrast, a MKlp2 mutant defective in binding myosin-II does not recruit Aurora B to the cell cortex and does not promote furrow formation during monopolar cytokinesis. This mutant is also defective in maintaining the ingressing furrow during bipolar cytokinesis. Together, these findings reveal that targeting Aurora B to the cell cortex (or the equatorial cortex) by MKlp2 is essential for the maintenance of the ingressing furrow for successful cytokinesis.
    PLoS ONE 06/2013; 8(6):e64826. DOI:10.1371/journal.pone.0064826 · 3.23 Impact Factor
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