Accurate inheritance of genomic content during cell division is dependent on synchronized changes in cellular organization and chromosome dynamics. In higher eukaryotes, once chromosomes have undergone condensation, the nuclear membranes, nuclear pore complexes, and nuclear lamina are disassembled in concert. The long-term goal of this project is to elucidate how events of mitosis are orchestrated. The interface between nuclear pore protein function and the cell cycle is an emerging and critical piece of this puzzle. Here, recently identified roles of the nucleoporin Nup153 at mitosis will be pursued. Mechanisms that integrate events of mitotic disassembly will also be investigated, as well as how disassembly is connected with nuclear reassembly at the end of mitosis. Specifically, in Aim 1, knockdown of Nup153 in mammalian cells in combination with a structure-function rescue analysis will be employed to decipher the functions of Nup153 in both early and late mitosis. In order to track which specific hallmarks of mitosis are altered, events of cell division will be monitored by live imaging. Identification of relevant protein partners, through both candidate and unbiased approaches, will be used to further hone in on the contributions of Nup153.
The second Aim i s focused on characterizing the role of sumoylation in regulating Nup153 function. The site(s) of sumoylation will be mapped and the functional consequence of interfering with this modification will be tested, both in knockdown-rescue experiments as well as with respect to specific features of pore architecture. Finally, in the third Aim, nuclei reconstituted in the cell-free Xenopus egg extract system will be used as a tool to investigate the role of the small GTPase Arf and its ability to stimulate phospholipase D as a node that provides an integrative signal for lamina and membrane disassembly. A role for COPI in organizing membrane populations at mitosis and how this affects the consequent participation of membranes in reassembly of the nuclear envelope will be tested. This research will yield new insight into the events of cell division and how they are coordinated. In turn, this information will impact our understanding of how abnormal nuclear morphology and DNA content arise, which is an imperative step in deciphering the molecular events that lead to cancerous cell proliferation.

Public Health Relevance

The goals of this research are to better understand how the events of cell division are orchestrated. When this coordination is disrupted, the cell nucleus does not reform properly. Such defects in morphology and DNA content are hallmarks of aggressive cancer cells, and the research proposed here will contribute to a better understanding of how these alterations arise.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM061275-13
Application #
8286293
Study Section
Special Emphasis Panel (ZRG1-CB-D (02))
Program Officer
Ainsztein, Alexandra M
Project Start
2000-07-01
Project End
2014-06-30
Budget Start
2012-07-01
Budget End
2014-06-30
Support Year
13
Fiscal Year
2012
Total Cost
$310,509
Indirect Cost
$102,811
Name
University of Utah
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
009095365
City
Salt Lake City
State
UT
Country
United States
Zip Code
84112
Chow, Kin-Hoe; Elgort, Suzanne; Dasso, Mary et al. (2014) The SUMO proteases SENP1 and SENP2 play a critical role in nucleoporin homeostasis and nuclear pore complex function. Mol Biol Cell 25:160-8
Shankaran, Sunita S; Mackay, Douglas R; Ullman, Katharine S (2013) A time-lapse imaging assay to study nuclear envelope breakdown. Methods Mol Biol 931:111-22
Chow, Kin-Hoe; Factor, Rachel E; Ullman, Katharine S (2012) The nuclear envelope environment and its cancer connections. Nat Rev Cancer 12:196-209
Powers, Matthew A; Fay, Marta M; Factor, Rachel E et al. (2011) Protein arginine methyltransferase 5 accelerates tumor growth by arginine methylation of the tumor suppressor programmed cell death 4. Cancer Res 71:5579-87
Mackay, Douglas R; Ullman, Katharine S; Rodesch, Christopher K (2010) Time-lapse imaging of mitosis after siRNA transfection. J Vis Exp :
Mackay, Douglas R; Makise, Masaki; Ullman, Katharine S (2010) Defects in nuclear pore assembly lead to activation of an Aurora B-mediated abscission checkpoint. J Cell Biol 191:923-31
Mackay, Douglas R; Elgort, Suzanne W; Ullman, Katharine S (2009) The nucleoporin Nup153 has separable roles in both early mitotic progression and the resolution of mitosis. Mol Biol Cell 20:1652-60
Lim, Roderick Y H; Ullman, Katharine S; Fahrenkrog, Birthe (2008) Biology and biophysics of the nuclear pore complex and its components. Int Rev Cell Mol Biol 267:299-342
Higa, Meda M; Alam, Steven L; Sundquist, Wesley I et al. (2007) Molecular characterization of the Ran-binding zinc finger domain of Nup153. J Biol Chem 282:17090-100
Ball, Jennifer R; Dimaano, Christian; Bilak, Amber et al. (2007) Sequence preference in RNA recognition by the nucleoporin Nup153. J Biol Chem 282:8734-40

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