The goal of our research is to determine the mechanisms by which the lamin isoform meshworks comprising the nuclear lamina (NL) organize chromatin in 3D and regulate gene expression. The experimental plan builds on recently developed techniques for determining lamin meshwork structures at nanoscale resolution using 3D-SIM combined with computational image analysis. This structural approach will be coordinated with studies of the biochemical/molecular interactions of each lamin isoform with chromatin using our recently developed HiLands model, which is the first to focus on the role of lamins in regulating chromatin organization. Throughout the proposed studies we will take advantage of our collection of single, double and triple lamin knockout mouse cells. Specifically we will elucidate the steps involved in the assembly of lamin isoform meshworks within the NL and determine how the assembly state of lamins impacts the 3D organization of chromatin and gene expression. These studies will be accompanied by a multi-pronged approach employing Hi-C, 4-C, epigenomics, transcriptomics, chromosome painting and FISH, the latter combined with 3D SIM- based immunostaining, to determine whether lamins regulate gene expression by maintaining 3D chromatin folding, enhancer-promoter interactions, and epigenetic modifications.

Public Health Relevance

The major goal of this research project is to determine the roles of the nuclear lamin proteins in organizing and regulating the genome. The lamins are major architectural proteins of the nucleus of mammalian cells where they are involved in the regulation of many nuclear processes. Interest in the importance of the lamins in regulating nuclear functions has increased in recent years due to the large number of mutations in lamin genes causing many diseases ranging from premature aging to cardiomyopathies and muscular dystrophies. The studies proposed in this grant use high-resolution techniques to determine how the nuclear structures formed by the lamins regulate the organization of the genome in both healthy and diseased cells.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM106023-06
Application #
9452993
Study Section
Special Emphasis Panel (ZRG1)
Program Officer
Flicker, Paula F
Project Start
2013-06-01
Project End
2020-12-31
Budget Start
2018-01-01
Budget End
2018-12-31
Support Year
6
Fiscal Year
2018
Total Cost
Indirect Cost
Name
Carnegie Institution of Washington, D.C.
Department
Type
DUNS #
072641707
City
Washington
State
DC
Country
United States
Zip Code
20005
Stephens, Andrew D; Liu, Patrick Z; Banigan, Edward J et al. (2018) Chromatin histone modifications and rigidity affect nuclear morphology independent of lamins. Mol Biol Cell 29:220-233
Huang, Yuejia; Li, Teng; Ems-McClung, Stephanie C et al. (2018) Aurora A activation in mitosis promoted by BuGZ. J Cell Biol 217:107-116
Zheng, Xiaobin; Hu, Jiabiao; Yue, Sibiao et al. (2018) Lamins Organize the Global Three-Dimensional Genome from the Nuclear Periphery. Mol Cell 71:802-815.e7
Gigante, Crystal M; Dibattista, Michele; Dong, Frederick N et al. (2017) Lamin B1 is required for mature neuron-specific gene expression during olfactory sensory neuron differentiation. Nat Commun 8:15098
Stephens, Andrew D; Banigan, Edward J; Adam, Stephen A et al. (2017) Chromatin and lamin A determine two different mechanical response regimes of the cell nucleus. Mol Biol Cell 28:1984-1996
Turgay, Yagmur; Eibauer, Matthias; Goldman, Anne E et al. (2017) The molecular architecture of lamins in somatic cells. Nature 543:261-264
Chen, Haiyang; Zheng, Xiaobin; Xiao, Danqing et al. (2016) Age-associated de-repression of retrotransposons in the Drosophila fat body, its potential cause and consequence. Aging Cell 15:542-52
Tran, Joseph R; Zheng, Xiaobin; Zheng, Yixian (2016) Lamin-B1 contributes to the proper timing of epicardial cell migration and function during embryonic heart development. Mol Biol Cell 27:3956-3963
West, Gun; Gullmets, Josef; Virtanen, Laura et al. (2016) Deleterious assembly of the lamin A/C mutant p.S143P causes ER stress in familial dilated cardiomyopathy. J Cell Sci 129:2732-43
Takeshi, Shimi; Pack, Chan-Gi; Goldman, Robert D (2016) Analyses of the Dynamic Properties of Nuclear Lamins by Fluorescence Recovery After Photobleaching (FRAP) and Fluorescence Correlation Spectroscopy (FCS). Methods Mol Biol 1411:99-111

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