Chromatin, the physical packaging of eukaryotic chromosomes, plays a major role in determining the patterns of gene silencing and expression across the genome. The active reorganization of chromatin structure, critical for gene regulation, is achieved by ATP-dependent machines called chromatin remodelers, which disassemble, slide, and reassemble nucleosomes on DNA. Disruptions in chromatin remodeler function perturb gene expression and have been directly linked with a number of cancers and developmental disorders. At present, it is not understood at a molecular level how remodelers reposition and reorganize nucleosomes, and what factors give rise to particular biochemical characteristics of remodelers. This proposal aims to uncover how different domains of the Chd1 remodeler participate in the nucleosome sliding reaction. X-ray crystallography will be used to visualize both the central ATPase motor as well as the C-terminal DNA-binding domain in complex with DNA substrates, which will reveal how remodelers recognize and distort duplex DNA. The contributions of the Chd1 DNA-binding domain to the speed, spacing, and direction of nucleosome sliding will be determined with Chd1 variants that have modified binding domains and/or variations in the linking segment to the ATPase motor. The ATPase motor is regulated by a pair of N-terminal chromodomains, which appear to enhance substrate specificity of the remodeler. Rapid kinetic analyses of nucleosome sliding reactions using stopped flow FRET will be used to identify stage(s) in the nucleosome sliding cycle affected by ATPase regulation. The results of this research will advance our understanding of how chromatin remodelers work and select their substrates, which are essential steps for interpreting changes in chromatin landscapes between healthy and diseased cells.

Public Health Relevance

The proper packaging of DNA into chromosomes is vitally important for normal cell growth and survival, and disruption of factors that organize chromosomes lead to many types of developmental diseases and cancer. Understanding how these organizing factors carry out their tasks is essential for identifying how cells are transformed to diseased states.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Project (R01)
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Macromolecular Structure and Function B Study Section (MSFB)
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Preusch, Peter
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Johns Hopkins University
Schools of Arts and Sciences
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Patel, Ashok; Chakravarthy, Srinivas; Morrone, Seamus et al. (2013) Decoupling nucleosome recognition from DNA binding dramatically alters the properties of the Chd1 chromatin remodeler. Nucleic Acids Res 41:1637-48
Patel, Ashok; McKnight, Jeffrey N; Genzor, Pavol et al. (2011) Identification of residues in chromodomain helicase DNA-binding protein 1 (Chd1) required for coupling ATP hydrolysis to nucleosome sliding. J Biol Chem 286:43984-93
Sharma, Amit; Jenkins, Katherine R; Heroux, Annie et al. (2011) Crystal structure of the chromodomain helicase DNA-binding protein 1 (Chd1) DNA-binding domain in complex with DNA. J Biol Chem 286:42099-104
McKnight, Jeffrey N; Jenkins, Katherine R; Nodelman, Ilana M et al. (2011) Extranucleosomal DNA binding directs nucleosome sliding by Chd1. Mol Cell Biol 31:4746-59
Hauk, Glenn; McKnight, Jeffrey N; Nodelman, Ilana M et al. (2010) The chromodomains of the Chd1 chromatin remodeler regulate DNA access to the ATPase motor. Mol Cell 39:711-23