How epigenetic states are inherited during S phase of the cell cycle is one of the most challenging questions in the chromatin and epigenetic fields. DNA replication-coupled nucleosome assembly plays an important role in epigenetic inheritance following DNA replication and DNA repair. Mutations of most, if not all, genes involved in replication-coupled nucleosome assembly result in defects in transcriptional silencing at heterochromatin and genome instability in both yeast and mammalian cells. We have been studying how nucleosomes are formed following DNA replication in yeast and human cells and have made multiple significant contributions to this process. However, how parental histone (H3-H4)2 tetramers are transferred to replicating DNA is still poorly understood, which hinders our understanding of transmission of epigenetic information into daughter cells. The major challenge to understanding parental histone (H3-H4)2 assembly is a lack of methods to track this process. Despite this challenge, we have developed the eSPAN (enrichment and Sequencing Protein- Associated Nascent DNA) method that can discern whether a protein binds to leading or lagging strands of DNA replication forks. This method enables us for the first time to monitor nucleosome assembly of both newly synthesized and parental histone (H3-H4)2 onto leading and lagging strands of DNA replication forks. In this proposal, we will elucidate molecular mechanisms whereby parental (H3-H4)2 are assembled into nucleosomes following DNA replication and how epigenetic marks are inherited during mitotic cell division in both yeast and human cells using a combination of genetic, biochemical and genomic approaches. Together, our studies should have a profound impact on the understanding of nucleosome assembly and epigenetic inheritance.

Public Health Relevance

Chromatin, an organized complex of DNA, RNA and proteins, encodes epigenetic information and maintains genome integrity. In recent years, it has become increasingly clear that alterations in chromatin states play a causal role in a variety of cancers. During S phase, chromatin structure is temporarily disrupted and disassembled in order for DNA replication to proceed. Following passage of the replication fork, distinct chromatin states marked by different histone modifications must be restored, which is a daunting challenge for cells. However, it is largely unknown how epigenetically determined chromatin states are inherited during S phase of the cell cycle. In this proposal, we will employ the novel method that we developed to detect whether a protein binds to leading or lagging strands of DNA replication forks, to study how parental (H3-H4)2 are assembled into nucleosomes, the 'first step' of inheritance of higher-order chromatin structure in both yeast and in human cells. These studies will not only provide an unprecedented insight into nucleosome assembly of parental (H3-H4)2, but also potentially increase our understanding of the contributions of epigenetic alterations to the development of human cancers.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Unknown (R35)
Project #
1R35GM118015-01
Application #
9067600
Study Section
Special Emphasis Panel (ZGM1-TRN-7 (MR))
Program Officer
Carter, Anthony D
Project Start
2016-09-01
Project End
2021-07-31
Budget Start
2016-09-01
Budget End
2017-07-31
Support Year
1
Fiscal Year
2016
Total Cost
$831,040
Indirect Cost
$311,640
Name
Columbia University (N.Y.)
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
621889815
City
New York
State
NY
Country
United States
Zip Code
10032
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Gan, Haiyun; Yu, Chuanhe; Devbhandari, Sujan et al. (2017) Checkpoint Kinase Rad53 Couples Leading- and Lagging-Strand DNA Synthesis under Replication Stress. Mol Cell 68:446-455.e3
Zhang, Kuo; Gao, Yuan; Li, Jingjing et al. (2016) A DNA binding winged helix domain in CAF-1 functions with PCNA to stabilize CAF-1 at replication forks. Nucleic Acids Res 44:5083-94