The PHD finger (Plant Homeodomain) module is a signature chromatin-associated protein motif. This module is present throughout eukaryotic proteomes, and mutations in the PHD fingers of many proteins are associated with cancers, immunodeficiency and mental retardation syndromes, and other genetic disorders. We previously demonstrated that a subset of PHD fingers act as high affinity binding modules for histone H3 trimethylated at lysine 4 (H3K4me3). We linked H3K4me3 to multiple different functions via its recognition by discrete PHD finger nuclear proteins, including providing the firs evidence that disrupting the read-out of a histone modification can cause an inherited human disease. Our long-term goal is to develop a comprehensive understanding of how PHD domain-containing proteins impact on chromatin dynamics and the relationship of such activities to fundamental nuclear functions and human disease processes. Here we focus on the multiple PHD domain-containing protein NSD2 (also named MMSET and WHSC1), a histone lysine methyltransferase implicated in the pathogenesis of the hematologic malignancy multiple myeloma. However, the molecular mechanism by which NSD2 regulates chromatin and the relationship of its enzymatic activity to disease pathogenesis is not well understood. Our preliminary work indicates that the primary physiologic activity at chromatin of NSD2 is dimethylation of histone H3 at lysine 36 (H3K36me2), and that NSD2 - via H3K36me2 catalysis - drives oncogenic programming in myeloma cells.
In Aim 1, we propose to extend our genomic studies and determine the genome-wide distribution of NSD2 in cancer and normal cells, and investigate the role of NSD2 activity and chromatin targeting in determining the H3K36me2 chromatin landscape. We also aim to elucidate the molecular mode of action for the NSD2 PHD domains and their role in the regulation of NSD2 cellular functions.
In Aim 2, we will characterize the mode of action for H3K36 methylation. We will identify proteins that preferentially recognize H3K36me2 and test the hypothesis that these proteins transduce NSD2 activity at chromatin to downstream biological outcomes. We will also explore the broader hypothesis that exquisite level of biological regulation can be achieved by subtle changes in histone methylation. The goal of Aim 3 is to identify new substrates of NSD2 using a novel chemical biological-proteomic strategy we have developed for proteome-wide discovery of functionally-relevant NSD2 substrates. The role of the most promising targets in regulation of nuclear pathways will be investigated using a combination of molecular and cellular approaches. These studies will identify new nuclear signaling pathways that are regulated by NSD2 and that may play a role in human disease. Together these studies will provide important new insights into how histone methylation regulates fundamental nuclear processes and the relationship of these activities to the pathogenesis of human diseases.

Public Health Relevance

We propose to investigate the molecular mode of action for the lysine methyltransferase and epigenetic regulator NSD2 in mammalian cells. Numerous human diseases, including cancer arise from epigenetic abnormalities. This proposal will provide new insight into how epigenetic mechanisms regulate important cellular functions, and has the potential to identify new targets for therapeutic intervention for human disease.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM079641-07
Application #
8598480
Study Section
Molecular Genetics B Study Section (MGB)
Program Officer
Carter, Anthony D
Project Start
2007-03-01
Project End
2016-12-31
Budget Start
2014-01-01
Budget End
2014-12-31
Support Year
7
Fiscal Year
2014
Total Cost
$282,600
Indirect Cost
$102,600
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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Carlson, Scott M; Gozani, Or (2014) Emerging technologies to map the protein methylome. J Mol Biol 426:3350-62
Wilkinson, Alex W; Gozani, Or (2014) Histone-binding domains: strategies for discovery and characterization. Biochim Biophys Acta 1839:669-75
Reynoird, Nicolas; Gozani, Or (2014) Nuclear PI5P, Uhrf1, and the road not taken. Mol Cell 54:901-3
Cheng, Zhongjun; Cheung, Peggie; Kuo, Alex J et al. (2014) A molecular threading mechanism underlies Jumonji lysine demethylase KDM2A regulation of methylated H3K36. Genes Dev 28:1758-71
Moore, Kaitlyn E; Carlson, Scott M; Camp, Nathan D et al. (2013) A general molecular affinity strategy for global detection and proteomic analysis of lysine methylation. Mol Cell 50:444-56
Paul, Shilpi; Kuo, Alex; Schalch, Thomas et al. (2013) Chd5 requires PHD-mediated histone 3 binding for tumor suppression. Cell Rep 3:92-102
Bua, Dennis J; Martin, Gloria Mas; Binda, Olivier et al. (2013) Nuclear phosphatidylinositol-5-phosphate regulates ING2 stability at discrete chromatin targets in response to DNA damage. Sci Rep 3:2137
Kuo, Alex J; Song, Jikui; Cheung, Peggie et al. (2012) The BAH domain of ORC1 links H4K20me2 to DNA replication licensing and Meier-Gorlin syndrome. Nature 484:115-9
Green, Erin M; Mas, Gloria; Young, Nicolas L et al. (2012) Methylation of H4 lysines 5, 8 and 12 by yeast Set5 calibrates chromatin stress responses. Nat Struct Mol Biol 19:361-3

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