The post-translational modification of histone proteins is now established as an important mechanism for regulating gene expression in eukaryotic cells. However, a structural and mechanistic understanding of how the histone modification enzymes function on their nucleosome substrate is lacking. This shortcoming limits interpretation of the wealth of genetic, genomic and biochemical data available, and it hampers development of new therapeutics that target the many chromatin enzymes associated with human diseases including cancer. To address this deficiency, we propose to study how the PRC1 polycomb repressive complex conjugates ubiquitin to histone H2A assembled in nucleosomes. The addition of ubiquitin to histone H2A has significant biomedical consequences: mono-ubiquitylation of histone H2A by PRC1 plays a critical role in embryonic stem cell maintenance, while mono-ubiquitylation of histone H2A by BRCA1 may mediate this breast cancer protein's tumor suppressor function. This proposal focuses on the following two specific aims: 1. Determine structure of the PRC1 ubiquitylation module on the nucleosome. We propose to crystallize the E2/E3 ubiquitylation complex of UbcH5c/Ring1/Bmi1 in complex with the nucleosome core particle and to determine the atomic structure of the complex by X-ray crystallography. 2. Elucidate mechanism for PRC1 module ubiquitylation of nucleosomes. The crystallographic studies will be complemented by biochemical studies to analyze the roles of the UbcH5c E2 ubiquitin-conjugating and the Ring1b/Bmi1 E3 ubiquitin ligase proteins in ubiquitylating nucleosomal H2A.

Public Health Relevance

In our cells, our DNA genetic information is packaged with histone proteins into chromatin. Recent studies have shown that histone proteins can be decorated by histone modification enzymes with small molecules or proteins, and these decorations regulate gene expression in normal and diseased cells. Our studies will visualize how the ubiquitin ligase class of histone modification enzymes associated with stem cells and cancer perform its function.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
1R01GM111651-01
Application #
8754079
Study Section
Special Emphasis Panel (ZRG1-BCMB-D (02))
Program Officer
Preusch, Peter
Project Start
2014-08-01
Project End
2018-07-31
Budget Start
2014-08-01
Budget End
2015-07-31
Support Year
1
Fiscal Year
2014
Total Cost
$280,091
Indirect Cost
$90,091
Name
Pennsylvania State University
Department
Biochemistry
Type
Schools of Arts and Sciences
DUNS #
003403953
City
University Park
State
PA
Country
United States
Zip Code
16802
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