The two specific aims in this proposal address fundamental mechanisms of regulating chromatin structure for transcription that are relevant to cancer. The SWI/SNF nucleosome-remodeling complex is present from yeast to humans where it functions as a tumor suppressor. We have found that the Snf2 ATPase subunit of the yeast complex is modified by acetylation, which causes its dissociation from acetylated histones. This is consistent with a model whereby acetylation of Snf2 disengages the SWI/SNF complex from target genes. In the first aim we will test this model in vitro and in vivo and further explore the consequences of Snf2 acetylation on SWI/SNF activities. Biochemical experiments will examine the effects of acetylating Snf2 on interaction of the complex with transcription factors, its recruitment to target genes, its function as an ATPase and its ability to remodel and displace promoter nucleosomes. In vivo studies will analyze the effect of Snf2 acetylation on SWI/SNF recruitment and function on candidate target genes. We will also perform genome wide analysis of the effect of Snf2 acetylation on the localization of the complex and its effect on gene expression. These studies will reveal fundamental information regarding the regulation of SWI/SNF activity, which will empower understanding its function as a tumor suppressor in humans. Acetylation of histone H3 on lysine 56 (AcK56) is enriched in several cancers and represents a mark of newly synthesized histones. Its presence in chromatin is indicative of incorporation of new histones into chromatin. The Set2 histone methyltransferase has been reported to function as a tumor suppressor in human cells and we have found that deletion of SET2 in yeast leads to the enrichment of H3AcK56 in transcribed regions. Set2 functions as part of the Set2/Rpd3 pathway by which RNA polymerase II signals for histone deacetylation during transcription. The fact that H3AcK56 accumulates in transcribed sequences in set2 mutations suggests that new histones replace the original histones in the absence of the Set2/Rpd3 pathway. In the second aim we will analyze the functions of Set2/Rpd3S pathway components in facilitating the retention of original histones during the passage of RNA polymerase II and preventing the insertion of H3AcK56 into transcribed chromatin. In addition we will test the role of the ISWI class of nucleosome remodelers in reassembling chromatin behind RNA polymerase II as we have found that they co-immunoprecipitate with Set2-methylated nucleosomes. These studies will reveal fundamental information regarding the process of reassembling chromatin behind RNA polymerase II, which is required to preserve the fidelity of transcription initiation. 1

Public Health Relevance

The SWI/SNF protein complex functions as a tumor suppressor in humans and this project will examine the regulation of the activity of this complex when it is modified. Acetylated histone H3 accumulates in the chromosomes of many cancer cells and this project will investigate the process that prevents this from happening in genes. 1

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM047867-23
Application #
8691850
Study Section
Molecular Genetics C Study Section (MGC)
Program Officer
Carter, Anthony D
Project Start
1992-08-01
Project End
2015-07-31
Budget Start
2014-08-01
Budget End
2015-07-31
Support Year
23
Fiscal Year
2014
Total Cost
Indirect Cost
Name
Stowers Institute for Medical Research
Department
Type
DUNS #
City
Kansas City
State
MO
Country
United States
Zip Code
64110
Venkatesh, Swaminathan; Li, Hua; Gogol, Madelaine M et al. (2016) Selective suppression of antisense transcription by Set2-mediated H3K36 methylation. Nat Commun 7:13610
Venkatesh, Swaminathan; Workman, Jerry L (2015) Histone exchange, chromatin structure and the regulation of transcription. Nat Rev Mol Cell Biol 16:178-89
Dutta, Arnob; Gogol, Madelaine; Kim, Jeong-Hoon et al. (2014) Swi/Snf dynamics on stress-responsive genes is governed by competitive bromodomain interactions. Genes Dev 28:2314-30
Khan, Dilshad H; Gonzalez, Carolina; Cooper, Charlton et al. (2014) RNA-dependent dynamic histone acetylation regulates MCL1 alternative splicing. Nucleic Acids Res 42:1656-70
Becker, Peter B; Workman, Jerry L (2013) Nucleosome remodeling and epigenetics. Cold Spring Harb Perspect Biol 5:
Venkatesh, Swaminathan; Workman, Jerry L (2013) Set2 mediated H3 lysine 36 methylation: regulation of transcription elongation and implications in organismal development. Wiley Interdiscip Rev Dev Biol 2:685-700
Smolle, Michaela; Workman, Jerry L (2013) Transcription-associated histone modifications and cryptic transcription. Biochim Biophys Acta 1829:84-97
Smolle, Michaela; Workman, Jerry L; Venkatesh, Swaminathan (2013) reSETting chromatin during transcription elongation. Epigenetics 8:10-5
Lin, Chia-Hui; Paulson, Ariel; Abmayr, Susan M et al. (2012) HP1a targets the Drosophila KDM4A demethylase to a subset of heterochromatic genes to regulate H3K36me3 levels. PLoS One 7:e39758
Smolle, Michaela; Venkatesh, Swaminathan; Gogol, Madelaine M et al. (2012) Chromatin remodelers Isw1 and Chd1 maintain chromatin structure during transcription by preventing histone exchange. Nat Struct Mol Biol 19:884-92

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