Shuster Charles B.

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Charles B.

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  • Article
    A global, myosin light chain kinase-dependent increase in myosin II contractility accompanies the metaphase–anaphase transition in sea urchin eggs
    (American Society for Cell Biology, 2006-07-12) Lucero, Amy ; Stack, Christianna ; Bresnick, Anne R. ; Shuster, Charles B.
    Myosin II is the force-generating motor for cytokinesis, and although it is accepted that myosin contractility is greatest at the cell equator, the temporal and spatial cues that direct equatorial contractility are not known. Dividing sea urchin eggs were placed under compression to study myosin II-based contractile dynamics, and cells manipulated in this manner underwent an abrupt, global increase in cortical contractility concomitant with the metaphase–anaphase transition, followed by a brief relaxation and the onset of furrowing. Prefurrow cortical contractility both preceded and was independent of astral microtubule elongation, suggesting that the initial activation of myosin II preceded cleavage plane specification. The initial rise in contractility required myosin light chain kinase but not Rho-kinase, but both signaling pathways were required for successful cytokinesis. Last, mobilization of intracellular calcium during metaphase induced a contractile response, suggesting that calcium transients may be partially responsible for the timing of this initial contractile event. Together, these findings suggest that myosin II-based contractility is initiated at the metaphase–anaphase transition by Ca2+-dependent myosin light chain kinase (MLCK) activity and is maintained through cytokinesis by both MLCK- and Rho-dependent signaling. Moreover, the signals that initiate myosin II contractility respond to specific cell cycle transitions independently of the microtubule-dependent cleavage stimulus.
  • Article
    Central spindle self-organization and cytokinesis in artificially activated sea urchin eggs
    (Marine Biological Laboratory, 2016-04) Henson, John H. ; Buckley, Mary W. ; Yeterian, Mesrob ; Weeks, Richard M. ; Simerly, Calvin R. ; Shuster, Charles B.
    The ability of microtubules of the mitotic apparatus to control the positioning and initiation of the cleavage furrow during cytokinesis was first established from studies on early echinoderm embryos. However, the identity of the microtubule population that imparts cytokinetic signaling is unclear. The two main––and not necessarily mutually exclusive–– candidates are the central spindle and the astral rays. In the present study, we examined cytokinesis in ammonia-activated sea urchin eggs, which lack paternally derived centrosomes and undergo mitosis mediated by unusual anastral, bipolar mini-spindles. Live cell imaging and immunolabeling for microtubules and the centralspindlin constituent and kinesin-related protein, MKLP1, demonstrated that furrowing in ammonia-activated eggs was associated with aligned arrays of centralspindlin-linked, opposed bundles of antiparallel microtubules. These autonomous, zipper- like arrays were not associated with a mitotic apparatus, but did possess characteristics similar to the central spindle region of control, fertilized embryos. Our results highlight the self-organizing nature of the central spindle region and its ability to induce cytokinesis-like furrowing, even in the absence of a complete mitotic apparatus.
  • Article
    Arp2/3 complex inhibition radically alters lamellipodial actin architecture, suspended cell shape, and the cell spreading process
    (American Society for Cell Biology, 2015-01-07) Henson, John H. ; Yeterian, Mesrob ; Weeks, Richard M. ; Medrano, Angela E. ; Brown, Briana L. ; Geist, Heather L. ; Pais, Mollyann D. ; Oldenbourg, Rudolf ; Shuster, Charles B.
    Recent studies have investigated the dendritic actin cytoskeleton of the cell edge's lamellipodial (LP) region by experimentally decreasing the activity of the actin filament nucleator and branch former, the Arp2/3 complex. Here we extend these studies via pharmacological inhibition of the Arp2/3 complex in sea urchin coelomocytes, cells that possess an unusually broad LP region and display correspondingly exaggerated centripetal flow. Using light and electron microscopy, we demonstrate that Arp2/3 complex inhibition via the drug CK666 dramatically altered LP actin architecture, slowed centripetal flow, drove a lamellipodial-to-filopodial shape change in suspended cells, and induced a novel actin structural organization during cell spreading. A general feature of the CK666 phenotype in coelomocytes was transverse actin arcs, and arc generation was arrested by a formin inhibitor. We also demonstrate that CK666 treatment produces actin arcs in other cells with broad LP regions, namely fish keratocytes and Drosophila S2 cells. We hypothesize that the actin arcs made visible by Arp2/3 complex inhibition in coelomocytes may represent an exaggerated manifestation of the elongate mother filaments that could possibly serve as the scaffold for the production of the dendritic actin network.