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Aurora-A Kinase and Inhibitor-2 Regulate the Cyclin Threshold for Mitotic Entry in Xenopus Early Embryonic Cell Cycles
David L. Satinover, David L. Brautigan and P. Todd Stukenberg
volume 5 | issue 19
1 october 2006Pages: 2268 - 2274
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The abrupt activation of CDK1 during mitotic entry requires suppression of CDK activity until a threshold concentration of cyclin B is synthesized, triggering the activation of a large pool of CDK. The cellular mechanisms that define the concentration of cyclin B at which the threshold occurs are unknown. Here we demonstrate that this threshold is regulated by Aurora-A kinase and phosphatase Inhibitor-2. In Xenopus CSF extracts that actively translate cyclin B1, immunodepletion of either endogenous xInhibitor-2 or endogenous xAurora-A caused delayed mitotic entry and normal timing was restored by addition of the respective recombinant proteins. Aurora-A depleted extracts also could be rescued by the addition of full-length xInhibitor-2, but not an xInhibitor-2 truncated of its PP1 binding motif. This demonstrates that inhibition of PP1 was required to compensate for the absence of Aurora-A. To test the hypothesis that the delays in mitotic entry in CSF extracts were due to increases in cyclin B thresholds, we employed interphase extracts, which are driven into mitosis by the addition of recombinant cyclin B in a non-linear (threshold) dose-response. Neutralization of endogenous xInhibitor-2 or xAurora with antibodies increased the cyclin B threshold concentration. Alternatively, the addition of exogenous Aurora-A or Inhibitor-2 lowered the concentration of cyclin B that triggered CDK activation. Because the cyclin B threshold could be raised or lowered by changing the amount of either Aurora-A or Inhibitor-2, the results demonstrate these regulatory proteins are involved in a signaling loop required to create the switching behavior characteristic of mitotic entry.
Authors
David L. Satinover
University of Virginia School of Medicine Charlottesville
David L. Brautigan
University of Virginia School of Medicine Charlottesville
P. Todd Stukenberg
University of Virginia School of Medicine Charlottesville
We now provide open access to journal articles published online for one year or more. This article may be downloaded at the following link:
Download PDF If the document does not open, please right-click on the link (control-click on a Macintosh) and select the option to save the file to disk.