Virtually all cells position their nucleus in specific locations that reflect cell and tissue function. Disruption of proteins involved in nuclear positioning pathways leads to altered cell physiology and human diseases including muscular dystrophies, cardiomyopathy, lissencephaly, cerebellar ataxia and dystonia. How nuclear positioning contributes to cell function is unclear. Our goal is to explore the mechanisms and function of nuclear position in migrating cells where nuclei are consistently positioned in the rea of cells. We have developed a model wounded monolayer system where nuclear movement establish this rearward position and can stimulated by external factors and measured quantitatively. We will also developed a novel method of artificially displacing nuclei in adherent cells by centrifugal force and will extend our studies to 3D migration systems where nuclear movements may be rate limiting for migration. Our earlier studies established that rearward nuclear position is actin- and myosin-dependent and requires the establishment of a connection between actin cables and the nucleus. This connection is mediated by the clustering of LINC complex components nesprin-2G in the outer nuclear membrane and SUN2 in the inner nuclear membrane. This clustering results in the formation of structures that resemble nuclear adhesions that we termed TAN lines for transmembrane actin-associated nuclear lines. We will further explore the assemble and function of TAN lines by pursuing exciting preliminary studies that suggest that in addition to mechanically coupling nuclei to the actin cytoskeleton, TAN lines exhibit mechanochemical signaling that affects nuclear movement. With our new method for artificially displacing nuclei, we have found surprising evidence that cells position their nuclei y actin or microtubule mechanisms depending on whether nuclei are displaced to the front of rear of the cells. We will use this system to define how a novel LINC complex associate with microtubules and microtubule motors and test whether cells switch between LINC complexes during polarization and active phases of cell migration. To understand how nuclear position influences migration, we will test the hypothesis that nuclei are coupled to focal adhesions and modulate their dynamics by an internal tension based mechanism. Lastly, we will explore the contribution of different LINC complexes to cell migration in 3D matrices and test the specific hypothesis that different LINC complexes may be required when nuclei deform to squeeze through small pores in the matrix. These studies will uncover novel mechanisms of LINC complex positioning of nuclei and provide new insights into how nuclear positioning influences cell migration. This information will contribute to understanding of how diseases associated with nuclear positioning pathways originate so that effective treatments can be designed.

Public Health Relevance

Cell migration is a fundamental property of most cells and contributes to development, wound healing, immune response and a host of other processes. This project is focused on understanding how the positioning of the nucleus in migrating cells is determined and how this influences the cell's ability to migrate. The focus is on molecules in the nuclear membrane that connect the nucleus to the cell's cytoskeletal system. These proteins are mutated in a number of human diseases, including muscular dystrophy, cardiomyopathy, lissencephaly, cerebellar ataxia and dystonia. Deciphering how these proteins function in nuclear positioning will contribute to understanding how these proteins cause disease and may provide insights to develop new therapies to treat these diseases.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM099481-08
Application #
9624778
Study Section
Nuclear and Cytoplasmic Structure/Function and Dynamics Study Section (NCSD)
Program Officer
Deatherage, James F
Project Start
2011-09-15
Project End
2020-01-31
Budget Start
2019-02-01
Budget End
2020-01-31
Support Year
8
Fiscal Year
2019
Total Cost
Indirect Cost
Name
Columbia University (N.Y.)
Department
Pathology
Type
Schools of Medicine
DUNS #
621889815
City
New York
State
NY
Country
United States
Zip Code
10032
Antoku, Susumu; Gundersen, Gregg G (2018) Analysis of Nesprin-2 Interaction with Its Binding Partners and Actin. Methods Mol Biol 1840:35-43
Lele, Tanmay P; Dickinson, Richard B; Gundersen, Gregg G (2018) Mechanical principles of nuclear shaping and positioning. J Cell Biol 217:3330-3342
Schrank, Benjamin R; Aparicio, Tomas; Li, Yinyin et al. (2018) Nuclear ARP2/3 drives DNA break clustering for homology-directed repair. Nature 559:61-66
Zhu, Ruijun; Liu, Chenshu; Gundersen, Gregg G (2018) Nuclear positioning in migrating fibroblasts. Semin Cell Dev Biol 82:41-50
Zhu, Ruijun; Gundersen, Gregg G (2018) Centrifugal Displacement of Nuclei in Adherent Cells to Study LINC Complex-Dependent Mechanisms of Homeostatic Nuclear Positioning. Methods Mol Biol 1840:91-100
Saunders, Cosmo A; Harris, Nathan J; Willey, Patrick T et al. (2017) TorsinA controls TAN line assembly and the retrograde flow of dorsal perinuclear actin cables during rearward nuclear movement. J Cell Biol 216:657-674
Zhu, Ruijun; Antoku, Susumu; Gundersen, Gregg G (2017) Centrifugal Displacement of Nuclei Reveals Multiple LINC Complex Mechanisms for Homeostatic Nuclear Positioning. Curr Biol 27:3097-3110.e5
Kawamura, Akira; Washington, Ilyas; Mihai, Doina M et al. (2017) Identification of ginkgolide targets in brain by photoaffinity labeling. Chem Biol Drug Des 89:475-481
Chang, Wakam; Gundersen, Gregg G (2016) Swollen Nuclei Signal from the Grave. Cell 165:1051-1052
Nader, Guilherme P F; Ezratty, Ellen J; Gundersen, Gregg G (2016) FAK, talin and PIPKI? regulate endocytosed integrin activation to polarize focal adhesion assembly. Nat Cell Biol 18:491-503

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