Structure-function studies of drosophila Mip120
Abstract/Contents
- Abstract
- The Drosophila dMyb-MuvB (MMB)/Drosophila RBF, dE2F2, and dMyb- interacting proteins (dREAM) is a multi-protein transcriptional regulator that regulates many biological processes including the cell cycle, development, and differentiation. MMB/dREAM contains multiple members that have DNA binding domains: dMyb, dE2F2-dDP, and Mip120. It is currently unclear which of these members can localize the rest of the complex to DNA. My studies focused on Mip120/LIN54. We wanted to determine which domains of Mip120 were necessary for its function, because previous studies of dMyb unexpectedly showed that the highly conserved dMyb DNA binding domain was not necessary to rescue Myb null lethality. In addition, we also wanted to determine which domains of Mip120 were sufficient for nuclear localization in different Drosophila cell types, including the egg chambers, wing discs, salivary glands, and eye discs. mip120 null females were sterile, had an ovary development phenotype, and did not lay eggs. Using the ovary development phenotype and egg laying defect as an assay for mip120 gene function, we conducted rescue experiments to determine which domains of Mip120 were necessary for its function. We found that multiple domains of Mip120, including the N-terminus, which contains two putative nuclear localization signals, and the C-terminus, which contains a highly conserved cysteine rich DNA binding domain (CXC) and a helix-coil-helix protein interaction domain (HCH) were necessary for mip120 function. From the nuclear localization experiments, we found that all of the Mip120 fragments were sufficient for nuclear localization. We found a previously unreported mip120 null phenotype. mip120 null nurse cell chromosomes were very condensed and had blobbier DNA than controls. The phenotype occurred very early in egg chamber development because even egg chambers as early as stage 2-4 chambers appeared blobbier in mip120 null mutants than controls. We attempted to determine the mechanism of mip120 null mutants. We found that global expression of many common epigenetic marks including H3K9me2/3, HP1, H3S10P, H3K4me3, and H4Ac remained similar in mip120 null mutants compared to controls. In mip120 null mutants, we also assayed which dREAM complex members still localized to DNA. Mip40 and dMyb no longer localized to DNA without Mip120, suggesting that Mip120 is necessary for dMyb and Mip40 nuclear and DNA localization. dE2F2 and p55 Caf1 still localized to DNA without Mip120, suggesting that dE2F2 and p55 Caf1 can localize to DNA independently of Mip120. Unexpectedly, Mip130 was also still able to localize to DNA without Mip120. Though Mip130 does not have a traditional DNA binding domain, it does have an AT hook, which may be able to localize Mip130 to the complex.
Description
| Type of resource | text |
|---|---|
| Form | electronic; electronic resource; remote |
| Extent | 1 online resource. |
| Publication date | 2015 |
| Issuance | monographic |
| Language | English |
Creators/Contributors
| Associated with | Cheng, Mei-Hsin |
|---|---|
| Associated with | Stanford University, Department of Genetics. |
| Primary advisor | Lipsick, Joseph Steven, 1955- |
| Thesis advisor | Lipsick, Joseph Steven, 1955- |
| Thesis advisor | Fuller, Margaret |
| Thesis advisor | Pollack, Jonathan D |
| Thesis advisor | Sage, Julien |
| Advisor | Fuller, Margaret |
| Advisor | Pollack, Jonathan D |
| Advisor | Sage, Julien |
Subjects
| Genre | Theses |
|---|
Bibliographic information
| Statement of responsibility | Mei-Hsin Cheng. |
|---|---|
| Note | Submitted to the Department of Genetics. |
| Thesis | Thesis (Ph.D.)--Stanford University, 2015. |
| Location | electronic resource |
Access conditions
- Copyright
- © 2015 by Mei-Hsin Cheng
- License
- This work is licensed under a Creative Commons Attribution Non Commercial 3.0 Unported license (CC BY-NC).
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