Genetic and biochemical interactions between the DNA replication initiator and a chromosome architecture protein in Bacillus subtilis
Thesis: Ph. D., Massachusetts Institute of Technology, Department of Biology, 2014.
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| Format: | Thesis |
| Language: | eng |
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Massachusetts Institute of Technology
2015
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| Online Access: | http://hdl.handle.net/1721.1/92594 |
| _version_ | 1811089814813933568 |
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| author | Seid, Charlotte A |
| author2 | Alan D. Grossman. |
| author_facet | Alan D. Grossman. Seid, Charlotte A |
| author_sort | Seid, Charlotte A |
| collection | MIT |
| description | Thesis: Ph. D., Massachusetts Institute of Technology, Department of Biology, 2014. |
| first_indexed | 2024-09-23T14:25:06Z |
| format | Thesis |
| id | mit-1721.1/92594 |
| institution | Massachusetts Institute of Technology |
| language | eng |
| last_indexed | 2024-09-23T14:25:06Z |
| publishDate | 2015 |
| publisher | Massachusetts Institute of Technology |
| record_format | dspace |
| spelling | mit-1721.1/925942019-04-12T12:25:14Z Genetic and biochemical interactions between the DNA replication initiator and a chromosome architecture protein in Bacillus subtilis Seid, Charlotte A Alan D. Grossman. Massachusetts Institute of Technology. Department of Biology. Massachusetts Institute of Technology. Department of Biology. Biology. Thesis: Ph. D., Massachusetts Institute of Technology, Department of Biology, 2014. This electronic version was submitted by the student author. The certified thesis is available in the Institute Archives and Special Collections. Cataloged from student-submitted PDF version of thesis. Includes bibliographical references (pages 204-228). I described the co-association, in vitro interaction, and epistatic relationship between the DNA replication initiator DnaA and the nucleoid-associated protein Rok in Bacillus subtilis. Using ChIP-seq, I characterized the genome-wide association of DnaA and its regulator YabA. I found that DnaA and YabA associated with over 30 chromosomal regions that were bound by Rok, and association of DnaA and YabA with these regions was dependent on Rok. The DNA-binding domain of DnaA was dispensable for association of DnaA with these chromosomal regions. This indirect Rok-dependent association contrasted with the canonical sequence-specific binding of DnaA to eight chromosomal regions containing clusters of DnaA boxes. I found that association of YabA with Rok-dependent regions did not require DnaA, in contrast to the DnaA-dependent association of YabA with DnaA box cluster regions. Furthermore, I showed an in vitro interaction between DnaA and Rok using purified proteins in an electrophoretic mobility shift assay. DnaA depended on Rok for association with a DNA probe, recapitulating the dependence observed at this chromosomal region in vivo. DnaA and Rok are both transcription factors and regulate some of the same genes. I analyzed global gene expression to determine the genetic relationship between these two transcriptional regulators. In general, rok was epistatic to dnaA; that is, the gene expression effects of DnaA depended on Rok, consistent with the ChIP-seq dependence. I investigated a potential role for Rok in regulating replication initiation, but I did not detect a replication phenotype of a rok null mutant under various conditions. I found that a rok null mutation did not detectably affect DnaA or YabA protein levels, and I showed that the stationary phase growth defect of this strain was dependent on comK, a downstream target of Rok. Additionally, I determined that a putative regulator of replication initiation, the pyruvate dehydrogenase enzyme PdhC, affected replication via its metabolic function but was not a direct regulator of replication initation. by Charlotte A. Seid. Ph. D. 2015-01-05T19:34:24Z 2015-01-05T19:34:24Z 2014 2014 Thesis http://hdl.handle.net/1721.1/92594 898127691 eng M.I.T. theses are protected by copyright. They may be viewed from this source for any purpose, but reproduction or distribution in any format is prohibited without written permission. See provided URL for inquiries about permission. http://dspace.mit.edu/handle/1721.1/7582 228 pages application/pdf Massachusetts Institute of Technology |
| spellingShingle | Biology. Seid, Charlotte A Genetic and biochemical interactions between the DNA replication initiator and a chromosome architecture protein in Bacillus subtilis |
| title | Genetic and biochemical interactions between the DNA replication initiator and a chromosome architecture protein in Bacillus subtilis |
| title_full | Genetic and biochemical interactions between the DNA replication initiator and a chromosome architecture protein in Bacillus subtilis |
| title_fullStr | Genetic and biochemical interactions between the DNA replication initiator and a chromosome architecture protein in Bacillus subtilis |
| title_full_unstemmed | Genetic and biochemical interactions between the DNA replication initiator and a chromosome architecture protein in Bacillus subtilis |
| title_short | Genetic and biochemical interactions between the DNA replication initiator and a chromosome architecture protein in Bacillus subtilis |
| title_sort | genetic and biochemical interactions between the dna replication initiator and a chromosome architecture protein in bacillus subtilis |
| topic | Biology. |
| url | http://hdl.handle.net/1721.1/92594 |
| work_keys_str_mv | AT seidcharlottea geneticandbiochemicalinteractionsbetweenthednareplicationinitiatorandachromosomearchitectureproteininbacillussubtilis |