The use of solid scaffolds that provide the correct mechanical and chemical cues to cells is a promising approach for guiding tissue repair, promoting tissue-scaffold integration and achieving adequate neovascularization. It is becoming increasingly clear that tuning scaffold rigidity is a powerful way to control cell function but how scaffold rigidity regulates gene expression is not well-understood. The focus of this proposal is on the molecular mechanisms by which gene expression is controlled by the mechanical properties of the substrate. We propose to test the hypothesis that substrate rigidity controls gene expression by tuning nuclear tension. Strong support for this hypothesis comes from our preliminary results: we have found that substrate rigidity significantly alters nuclear shape through the modulation of cytoskeletal forces. We have also established that cytoskeletal force transfer to the nuclear surface is mediated by nuclear membrane embedded LINC (for linker of nucleoskeleton to cytoskeleton) complex proteins. Our approach is to 1) determine which genes are turned on or off in a substrate rigidity dependent manner, 2) examine the extent to which LINC complex proteins are required for rigidity control of genes, and 3) characterize the mechanisms by which rigidity modulation of nuclear shape controls intra-nuclear chromatin structure, spatial location of genes and epigenetic modifications that collectively regulate gene expression.
Two specific aims are proposed:
Aim 1 : To test the hypothesis that substrate rigidity controls gene expression in a LINC complex dependent manner.
Aim 2 : To characterize the mechanisms by which nuclear tension regulates the expression of genes. The successful completion of these aims will have broad-ranging impact, in fields as diverse as cell-biomaterial interactions, nuclear and cell mechanics and molecular and cell biology of gene regulation. Collectively, this work is of strong interest to both engineering and scientific disciplines. The project integrates the expertise of three collaborators (Lele, Nickerson and Roux) from very different backgrounds (bioengineering, molecular biology, cell biology). The interaction between investigators of such varied background is expected to result in new and highly significant discoveries in the proposed problem area. Each investigator will contribute innovative, cutting-edge techniques in the fields of molecular biology, cell and molecular imaging, biomaterials and cell and nuclear mechanics. The completion of these aims will enhance our understanding of how scaffold properties direct vascular cells. As a result, we expect that they will promote the development of improved scaffolds for many tissue engineering applications.

Public Health Relevance

Many applications in tissue engineering involve cell culture on solid scaffolds with defined properties. We seek to improve scientific understanding of how scaffold properties regulate gene expression in cells. This will help improve our ability to contro cells and hence engineer tissues with superior performance.

Agency
National Institute of Health (NIH)
Institute
National Institute of Biomedical Imaging and Bioengineering (NIBIB)
Type
Research Project (R01)
Project #
5R01EB014869-03
Application #
8705518
Study Section
Biomaterials and Biointerfaces Study Section (BMBI)
Program Officer
Hunziker, Rosemarie
Project Start
2012-08-01
Project End
2016-07-31
Budget Start
2014-08-01
Budget End
2015-07-31
Support Year
3
Fiscal Year
2014
Total Cost
Indirect Cost
Name
University of Florida
Department
Engineering (All Types)
Type
Biomed Engr/Col Engr/Engr Sta
DUNS #
City
Gainesville
State
FL
Country
United States
Zip Code
32611
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Mehus, Aaron A; Anderson, Ruthellen H; Roux, Kyle J (2016) BioID Identification of Lamin-Associated Proteins. Methods Enzymol 569:3-22
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Alam, Samer G; Zhang, Qiao; Prasad, Nripesh et al. (2016) The mammalian LINC complex regulates genome transcriptional responses to substrate rigidity. Sci Rep 6:38063
Kim, Dae In; Roux, Kyle J (2016) Filling the Void: Proximity-Based Labeling of Proteins in Living Cells. Trends Cell Biol 26:804-817
Wu, Qiong; Sharma, Soni; Cui, Hang et al. (2016) Targeting the chromatin remodeling enzyme BRG1 increases the efficacy of chemotherapy drugs in breast cancer cells. Oncotarget 7:27158-75
Kent, Ian A; Lele, Tanmay P (2016) Microtubule-based force generation. Wiley Interdiscip Rev Nanomed Nanobiotechnol :
Aggarwal, Varun; Dickinson, Richard B; Lele, Tanmay P (2016) Concentration Sensing by the Moving Nucleus in Cell Fate Determination: A Computational Analysis. PLoS One 11:e0149213

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