Regulation and Dynamics of Cell Division

The lab exploits the genetics and cell biology of Drosophila to explore 2 aspects of mitosis:

1. The control of the metaphase-anaphase transition exerted by the spindle assembly checkpoint.
2. The regulation of nonmuscle myosin II by the mitotic cycle and its role in the earliest steps of Drosophila embryogenesis.

Our experimental approaches combine live cell imaging with the genetic tools of Drosophila to examine in vivo the dynamics and functions of key proteins during normal and perturbed mitosis.    

(1) The spindle assembly checkpoint (SAC).
During mitosis, chromosomes must attach correctly to the spindle so that chromatids will segregate accurately to opposite poles when the cell divides. Improperly attached kinetochores lead to aneuploidy with grave consequences for the daughter cells (Fig 1).  Such attachment errors are very rare in normal cells because of a surveillance system called the spindle checkpoint. This process detects improperly attached kinetochores during prometaphase, and generates a signal that inhibits the onset of anaphase, thus providing more time for correct attachements to form.

Fig 1: Normal and abnormal mitosis in Drosphila neuroblasts
Examples of mitosis showing metaphase (left), normal anaphase (top), and abnormal anaphase (bottom), the consequences of defective spindle function and defective checkpoint. Chromosomes are fixed and stained with aceto-orcein.

The molecular mechanism of the spindle checkpoint is the subject of intense study in many labs, including ours, in part because aneuploidy is implicated in the etiology of cancer.  Outstanding issues include the nature of the anaphase inhibitor, the mechanism by which the inhibitor is generated,  the roles of the different molecular components,  the mechanism by which the checkpoint is shut off once correct kinetochore linkages are formed. 

(2) Regulation of nonmuscle myosin II by the mitotic cycle.
The actin-myosin cytoskeleton is subjected to profound cell-cycle-dependent changes in organization and activity. Most dramatically, actin and myosin are transiently assembled into the cytokinetic contractile ring soon after the start of mitotic exit. Using the early Drosophila embryo as a model, and GFP-tagged nonmuscle (cytoplasmic) myosin II, we have shown that myosin activity is negatively controlled both spatially and temporally by the mitotic oscillations of Cdc2 kinase activity (Fig 2). We are currently using this as a model to better understand the regulation of cytokinesis.

Fig 2: Cortical GFP-Myosin undergoes cycles of activation and inactivatiopn linked to the mitotic cycle of the early syncytial embryo.
Confocal images of a living fly embryo during cycles 3-10.  Myosin is activated during interphase and recruited in a "patch" of cortex that grows with each mitotic cycle until the whole surface of the egg is involved.  This cyclical myosin activity is critical to the proper migration of the nuclei within the syncytium. See Royou et al 2002.

Selection of Publications

Royou, A., Gagou, M.E., Karess R.E., Sullivan W.  (2010) BubR1 and Polo-coated DNA tethers facilitate the poleward segregation of acentric chromosomes. Cell 140: 235-245.
Abstract

Rahmani, Z., Gagou, M.E.  Emre, D., Lefebvre,  C. , Karess, RE. (2009). Separating the spindle checkpoint and timer activities of BubR1.  J. Cell Biol. 187: 597-605
Abstract

Katsani, K., Karess, RE., Dostatni, N., Doye V (2008). In vivo dynamics of Drosophila nuclear envelope components Mol Biol. Cell 19:3652-3666.
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Buffin, E., Emre, D., Karess, R. (2007) Flies without a spindle checkpoint. Nature Cell Bio. 9:565-72.
Abstract

Buffin, E., Lefebvre, C., Huang, J-Y, Gagou, ME, Karess, R. (2005) Recruitment of Mad2 to the kinetochore requires the Rod/Zw10 complex. Current Biology 15: 856-861. 
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