Gene activation and silencing via epigenetic modifications is important for determining cell fate and is critical for the proper development of multicellular organisms. Determining the relative importance of various epigenetic markers in specific developmental processes can be challenging because these modifications are often dynamic and vary between cell and tissue types. Common approaches such as a gene knockout of an enzyme required for epigenetic modifications can be used to assess the global importance of the modification, but do not provide a straightforward approach for testing the importance of the modification at specific loci, cell types and developmental time points. Here, we aim to create a general approach for the control of epigenetic modifications that is reversible and can be applied at specific times in development to a specified set of cells. In preliminary studies we have developed a light activatable protein that localizes to the cytoplasm in the dark, but enters the nucleus when the cell is illuminated with blue light. Our hypothesis is that this switch, called LANS for Light Activatable Nuclear Shuttle, can be used to control the activity of proteins that must be in the nucleus to be functional. As proof of concept, we have shown that we can use LANS to control the activity of a transcription factor in yeast with light. Here, we wil explore if light activated nuclear localization can be used to control enzymes and scaffolding proteins required for histone and DNA methylation and acetylation. To target predetermined loci, we will fuse LANS with naturally occurring and engineered DNA binding domains. We will test the LANS switch in mammalian cell culture, and we will explore whether the switch can be used to manipulate cell fate in C. elegans via light mediated control of a histone deacetylase.

Public Health Relevance

We are developing new light activatable proteins that can be used to manipulate epigenetic modifications in specific cells at specific time points during the development of an organism. Dynamic changes to epigenome are critical for biologically important events such as gene expression and DNA repair, and play an important role in cancer development and stem cell biology. Our new tools for manipulating the epigenome will allow researchers to determine the relative importance of various epigenetic markers in normal and diseased cells.

Agency
National Institute of Health (NIH)
Institute
National Institute on Drug Abuse (NIDA)
Type
Research Project (R01)
Project #
1R01DA036877-01
Application #
8642354
Study Section
Special Emphasis Panel (ZRG1-BST-N (50))
Program Officer
Satterlee, John S
Project Start
2013-09-30
Project End
2018-05-31
Budget Start
2013-09-30
Budget End
2014-05-31
Support Year
1
Fiscal Year
2013
Total Cost
$397,248
Indirect Cost
$134,180
Name
University of North Carolina Chapel Hill
Department
Biochemistry
Type
Schools of Medicine
DUNS #
608195277
City
Chapel Hill
State
NC
Country
United States
Zip Code
27599
Rothbart, Scott B; Strahl, Brian D (2014) Interpreting the language of histone and DNA modifications. Biochim Biophys Acta 1839:627-43