Access to the Genome: A Study of Transcription Factor Binding Within Nucleosomes
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The Ohio State University / OhioLINK
2016
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| Online Access: | http://rave.ohiolink.edu/etdc/view?acc_num=osu1480603783786784 |
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ndltd-OhioLink-oai-etd.ohiolink.edu-osu14806037837867842021-08-03T06:39:22Z Access to the Genome: A Study of Transcription Factor Binding Within Nucleosomes Brehove, Matthew Steven Biochemistry Biophysics Physics Nucleosome Hexasome transcription factor single molecule single-molecule FRET PIFE Histone octamer TIRF Fluorescence Resonance Energy Transfer All the DNA in a cell’s nucleus is packaged into a material called chromatin consisting of DNA and DNA-associated proteins. The basic unit of chromatin is the nucleosome which consists of ~147 bases of DNA wrapped around a protein core composed of two copies each of histones H2A, H2B, H3, and H4. DNA wrapped into a nucleosome is inaccessible to most DNA processing machinery. This machinery needs access to the DNA to perform processes such as transcription, replication, and repair. The cell uses many mechanisms to modulate this protection including nucleosome unwrapping, sliding, remodeling, disassembly, and chemical modification. This work used transcription factors along with fluorescent methods to probe the accessibility of DNA near the ends of the nucleosome. The DNA in this entry-exit region can spontaneously unwrap to provide transient access to DNA binding factors such as transcription factors. We studied the effect of histone post-translational modifications on the accessibility of DNA in the entry-exit region. We found that the phosphorylation at H3Y41ph increased DNA accessibility in vitro by 3-fold, which is similar in size to the effect of a previously studied modification H3K56ac. Since both of these modifications are associated with active genes and could occur together, we studied nucleosomes with both modifications and they showed a 17±5-fold increase in accessibility. This indicates that the cell could use multiple modifications in the entry-exit region to adjust the accessibility of nucleosomal DNA by over an order of magnitude. We also studied the effects of the removal of one H2A/H2B dimer. These partially formed nucleosomes, called hexasomes, are a component of chromatin that has been largely unstudied. Our results have shown that hexasomes have DNA accessibility similar to that of fully-formed nucleosomes on the side with the remaining H2A/H2B dimer. However, on the side without the dimer, our results showed the DNA remains permanently unwrapped by approximately 40 bases. The unwrapping caused this dimer-distal side to have far greater DNA accessibility amounting to that of naked DNA. These results show that there is a dramatic alteration of DNA accessibility caused by the formation of hexasomes. 2016 English text The Ohio State University / OhioLINK http://rave.ohiolink.edu/etdc/view?acc_num=osu1480603783786784 http://rave.ohiolink.edu/etdc/view?acc_num=osu1480603783786784 unrestricted This thesis or dissertation is protected by copyright: all rights reserved. It may not be copied or redistributed beyond the terms of applicable copyright laws. |
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NDLTD |
| language |
English |
| sources |
NDLTD |
| topic |
Biochemistry Biophysics Physics Nucleosome Hexasome transcription factor single molecule single-molecule FRET PIFE Histone octamer TIRF Fluorescence Resonance Energy Transfer |
| spellingShingle |
Biochemistry Biophysics Physics Nucleosome Hexasome transcription factor single molecule single-molecule FRET PIFE Histone octamer TIRF Fluorescence Resonance Energy Transfer Brehove, Matthew Steven Access to the Genome: A Study of Transcription Factor Binding Within Nucleosomes |
| author |
Brehove, Matthew Steven |
| author_facet |
Brehove, Matthew Steven |
| author_sort |
Brehove, Matthew Steven |
| title |
Access to the Genome: A Study of Transcription Factor Binding Within Nucleosomes |
| title_short |
Access to the Genome: A Study of Transcription Factor Binding Within Nucleosomes |
| title_full |
Access to the Genome: A Study of Transcription Factor Binding Within Nucleosomes |
| title_fullStr |
Access to the Genome: A Study of Transcription Factor Binding Within Nucleosomes |
| title_full_unstemmed |
Access to the Genome: A Study of Transcription Factor Binding Within Nucleosomes |
| title_sort |
access to the genome: a study of transcription factor binding within nucleosomes |
| publisher |
The Ohio State University / OhioLINK |
| publishDate |
2016 |
| url |
http://rave.ohiolink.edu/etdc/view?acc_num=osu1480603783786784 |
| work_keys_str_mv |
AT brehovematthewsteven accesstothegenomeastudyoftranscriptionfactorbindingwithinnucleosomes |
| _version_ |
1719440995368566784 |