Regulation of eukaryotic gene expression at the transcriptional level relies on the coordinated action of multiple protein factors. The recruitment of multi-subunit coregulator complexes containing chromatin-modifying activities by sequence-specific DNA binding factors and chromatin-binding proteins to specific loci on the genome is an important step in regulating gene transcription. The Sin3 corepressor complex is one of only a handful of major corepressor complexes identified thus far in mammalian cells. The negative influence on gene transcription exerted by this complex relies in part on the action of nuclear histone deacetylases (HDACs), a group of enzymes that catalyze the hydrolysis of acetyl groups from acetylated chromatin substrates. The ~150 kDa Sin3 protein functions both as a molecular scaffold for corepressor complex assembly and interacts directly, or indirectly through other components of the complex, with a surprisingly large and diverse group of DNA-binding transcription factors and chromatin-binding proteins that play important roles in normal growth, development, and cellular homeostasis. Our studies for the next funding period will address questions broadly relating to how components of the Sin3 corepressor complex are recruited by diverse promoter-bound transcription factors on the one hand and by specific patterns of post-translationally modified histones, also referred to as """"""""chromatin marks,"""""""" on the other. Another major question relates to how the complex is assembled and how it contributes to its functional diversity. Answers to these questions rely on structure-function analyses. The primary tool for the structural investigations will be solution-state NMR spectroscopy because it is ideally suited for studies of transcription factors as they frequently harbor natively unfolded segments and also because of the relatively small size and tractable nature of the systems being studied. In addition, the technique is well-suited for studies of weak interactions such as those involving chromatin-binding modules. Functional analyses will rely on a variety of biochemical and biophysical approaches.
The aims of the project broadly focus on three themes: (1) recruitment of the Sin3 corepressor by promoter-bound transcription factors, (2) recruitment and targeting of the corepressor complex to specific chromatin marks and (3) assembly of the core corepressor complex.

Public Health Relevance

The transcription of eukaryotic genes into messenger RNAs for subsequent translation to produce proteins is regulated by a class of proteins known as transcriptional coregulators. These coregulators are recruited to specific regions of the genome by proteins that bind DNA specifically and/or by proteins that bind histones. The molecular interactions involving the Sin3 coregulator and those involving histone-binding proteins that interact with Sin3 are the focus of this proposal because little is known about how these proteins function and because defects in and abnormal amounts of these proteins are associated with diseases such as cancer. The proposed work will attempt to derive detailed molecular pictures of proteins captured in the act of interacting with Sin3 or those of Sin3-interacting proteins captured in the act of interacting with histones, attempt to rationalize how these interactions or lack thereof could lead to diseased states, and provide ideas for developing suitable molecular therapies.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM064715-08
Application #
7858295
Study Section
Macromolecular Structure and Function C Study Section (MSFC)
Program Officer
Preusch, Peter C
Project Start
2002-04-01
Project End
2012-05-31
Budget Start
2010-06-01
Budget End
2011-05-31
Support Year
8
Fiscal Year
2010
Total Cost
$285,990
Indirect Cost
Name
Northwestern University at Chicago
Department
Biochemistry
Type
Schools of Arts and Sciences
DUNS #
160079455
City
Evanston
State
IL
Country
United States
Zip Code
60201
Xie, Tao; Zmyslowski, Adam M; Zhang, Yongbo et al. (2015) Structural Basis for Multi-specificity of MRG Domains. Structure 23:1049-57
Clark, Michael D; Marcum, Ryan; Graveline, Richard et al. (2015) Structural insights into the assembly of the histone deacetylase-associated Sin3L/Rpd3L corepressor complex. Proc Natl Acad Sci U S A 112:E3669-78
Xie, Tao; Graveline, Richard; Kumar, Ganesan Senthil et al. (2012) Structural basis for molecular interactions involving MRG domains: implications in chromatin biology. Structure 20:151-60
Kumar, Ganesan Senthil; Chang, William; Xie, Tao et al. (2012) Sequence requirements for combinatorial recognition of histone H3 by the MRG15 and Pf1 subunits of the Rpd3S/Sin3S corepressor complex. J Mol Biol 422:519-31
Xie, Tao; He, Yuan; Korkeamaki, Hanna et al. (2011) Structure of the 30-kDa Sin3-associated protein (SAP30) in complex with the mammalian Sin3A corepressor and its role in nucleic acid binding. J Biol Chem 286:27814-24
Kumar, Ganesan Senthil; Xie, Tao; Zhang, Yongbo et al. (2011) Solution structure of the mSin3A PAH2-Pf1 SID1 complex: a Mad1/Mxd1-like interaction disrupted by MRG15 in the Rpd3S/Sin3S complex. J Mol Biol 408:987-1000
He, Yuan; Imhoff, Rebecca; Sahu, Anirban et al. (2009) Solution structure of a novel zinc finger motif in the SAP30 polypeptide of the Sin3 corepressor complex and its potential role in nucleic acid recognition. Nucleic Acids Res 37:2142-52
Sahu, Sarata C; Swanson, Kurt A; Kang, Richard S et al. (2008) Conserved themes in target recognition by the PAH1 and PAH2 domains of the Sin3 transcriptional corepressor. J Mol Biol 375:1444-56
He, Yuan; Radhakrishnan, Ishwar (2008) Solution NMR studies of apo-mSin3A and -mSin3B reveal that the PAH1 and PAH2 domains are structurally independent. Protein Sci 17:171-5
Zhang, Yongbo; Zhang, Zhipeng; Demeler, Borries et al. (2006) Coupled unfolding and dimerization by the PAH2 domain of the mammalian Sin3A corepressor. J Mol Biol 360:7-14

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