The central problems addressed in the proposed research are how the centrosome is assembled, how it duplicates once per cell cycle, and what happens in cells in which centrosome number is aberrant. The centrosome nucleates microtubules and organizes those microtubules to create a useful array. The centrosome is the major microtubule organizing center in animal cells, and is present in a single copy at the beginning of the cell cycle. The centrosome duplicates in S phase, and the two resulting centrosomes help to organize the two poles of the mitotic spindle. Work from my lab and others over the last ten years has identified molecules involved in microtubule nucleation, the central regulators of centrosome duplication, important structural proteins involved in duplication, and control mechanisms that control centrosome number and link the centrosome and the cell cycle. Interest in the centrosome has grown recently because a correlation has been established between centrosome abnormalities and the development of cancer. Cancer cells often have extra centrosomes, which is likely to contribute to the genomic instability and rapid evolution characteristic of this disease. The proposed experiments make use of the strengths that we have developed in reagents and assays for studying centrosome structure, function and duplication. I address four specific aims in this proposal: 1) Characterize the block to centrosome reduplication. We will test models for the mechanism of the block, determine whether cancer cells have defects in the block, and test whether the block can be overcome by manipulation of potential regulators. 2) Investigate the relationship between centrosome number, ploidv, and genome instability. We will use cell fusion methods to create cells of defined centrosome number and ploidy, examine the progression and outcome of mitosis in these cells, and compare normal and cancer cells in their response to centrosome and ploidy abnormalities. 3) Define the molecular interactions of the gamma-tubulin ring complex with microtubules and with the centrosome. We will use purified components to determine the molecular interactions between the gamma-tubulin ring complex, the microtubule, and the centrosome. 4) Investigate the role of delta-tubulin and epsilon-tubulin in centrosome function and duplication. We will characterize the interactions between delta-tubulin, epsilon-tubulin and the other known tubulins, and define the function of delta-tubulin in vitro using assays in frog egg extracts, and in vivo in human cells.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
2R01GM052022-10
Application #
6928858
Study Section
Cell Development and Function Integrated Review Group (CDF)
Program Officer
Deatherage, James F
Project Start
1995-08-01
Project End
2009-03-31
Budget Start
2005-04-01
Budget End
2006-03-31
Support Year
10
Fiscal Year
2005
Total Cost
$374,827
Indirect Cost
Name
Stanford University
Department
Biology
Type
Schools of Arts and Sciences
DUNS #
009214214
City
Stanford
State
CA
Country
United States
Zip Code
94305
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Vladar, Eszter K; Lee, Yin Loon; Stearns, Tim et al. (2015) Observing planar cell polarity in multiciliated mouse airway epithelial cells. Methods Cell Biol 127:37-54
Firat-Karalar, Elif Nur; Stearns, Tim (2015) Probing mammalian centrosome structure using BioID proximity-dependent biotinylation. Methods Cell Biol 129:153-170
Van de Mark, Daniel; Kong, Dong; Loncarek, Jadranka et al. (2015) MDM1 is a microtubule-binding protein that negatively regulates centriole duplication. Mol Biol Cell 26:3788-802
Turk, Erin; Wills, Airon A; Kwon, Taejoon et al. (2015) Zeta-Tubulin Is a Member of a Conserved Tubulin Module and Is a Component of the Centriolar Basal Foot in Multiciliated Cells. Curr Biol 25:2177-83
Lee, Yin Loon; Santé, Joshua; Comerci, Colin J et al. (2014) Cby1 promotes Ahi1 recruitment to a ring-shaped domain at the centriole-cilium interface and facilitates proper cilium formation and function. Mol Biol Cell 25:2919-33
Firat-Karalar, Elif Nur; Rauniyar, Navin; Yates 3rd, John R et al. (2014) Proximity interactions among centrosome components identify regulators of centriole duplication. Curr Biol 24:664-70
Lee, Joanna Y; Hong, Wan-Jen; Majeti, Ravindra et al. (2014) Centrosome-kinase fusions promote oncogenic signaling and disrupt centrosome function in myeloproliferative neoplasms. PLoS One 9:e92641
Firat-Karalar, Elif N; Sante, Joshua; Elliott, Sarah et al. (2014) Proteomic analysis of mammalian sperm cells identifies new components of the centrosome. J Cell Sci 127:4128-33
Stearns, Tim (2014) Journey to the center of the centrosome. Dev Cell 28:603-4

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