This research project is to identify FoxA1 interacting partners. In this study, a potential FoxA1 binding partner, Upstream Stimulatory Factor 2 (USF2), has been examined; its DNA binding activity was determined and the function of USF2 was monitored in USF2 knockdown LNCaP cells. FLAG-6His-FoxA1 stable expressing LNCaP cell lines were established. The FoxA1 protein complex from LNCaP tagged-FoxA1 cells was purified and analyzed by Liquid Chromatography Tandem Mass Spectrometry.
USF2 has been confirmed to interact with FoxA1 using ImmunoPrecipitation and glutathione-S-transferase pull-down assays. Our study also indicates, as FoxA1, USF2 binds to prostatic specific gene promoters such as probasin promoter, Spermine Binding Protein promoter, and the Prostate Specific Antigen core enhancer. PSA mRNA levels were increased in USF2 knocked down LNCaP cells compared with control LNCaP cells when the LNCaP cells were treated with DHT (10-8 M).
Using LC-MS/MS analysis, a total of 16 proteins that appear to interact with Foxa1 have been identified and eight of them have been previously reported as AR interacting proteins. Further work is needed to confirm the binding between FoxA1 and these identified proteins and to perform functional studies of the confirmed FoxA1 binding partners.
This study has identified FoxA1 interacting partners which helps unravel the critical transcription factors that control cell type specific gene expression and to further understand the role of FoxA1 during prostate development. This work gives us an insight into the essential mechanism of androgen regulation of the prostate gland and of hormonal regulation of prostate cancer.
| Identifer | oai:union.ndltd.org:VANDERBILT/oai:VANDERBILTETD:etd-04132010-231904 |
| Date | 17 April 2010 |
| Creators | SUN, QIAN |
| Contributors | Sarki Abba Abdulkadir, Robert J. Matusik, Jennifer A. Pietenpol, David L. Hachey |
| Publisher | VANDERBILT |
| Source Sets | Vanderbilt University Theses |
| Language | English |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | http://etd.library.vanderbilt.edu/available/etd-04132010-231904/ |
| Rights | unrestricted, I hereby certify that, if appropriate, I have obtained and attached hereto a written permission statement from the owner(s) of each third party copyrighted matter to be included in my thesis, dissertation, or project report, allowing distribution as specified below. I certify that the version I submitted is the same as that approved by my advisory committee. I hereby grant to Vanderbilt University or its agents the non-exclusive license to archive and make accessible, under the conditions specified below, my thesis, dissertation, or project report in whole or in part in all forms of media, now or hereafter known. I retain all other ownership rights to the copyright of the thesis, dissertation or project report. I also retain the right to use in future works (such as articles or books) all or part of this thesis, dissertation, or project report. |
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