These investigations focus on the regulation of alpha-fetoprotein (AFP) gene expression as a tumor-associated gene switch. During liver regeneration, hepatocarcinoma or germinal tumor generation, AFP transcription is specifically activated. Though widely applied as a diagnostic marker, the regulatory control mechanisms involved in tumorigenic expression of AFP remain largely undetermined, as does the importance of this reactivation for prognostic and therapeutic value.
The specific aims of these proposed studies are as follows:
Specific Aim 1 : To determine the DNA domain(s) that mediates aberrant reactivation of AFP gene expression by: a) construction of a series of AFP deletion and hybrid gene insertion templates, and b) employing these templates in cell culture and transcription systems that reconstitute adult liver repression and hepatoma activation of AFP expression in vitro.
Specific Aim 2 : To model the stagewise progression of tumorigenesis by activating AFP gene expression within synthetic nuclei. We will undertake this by: a) assembly of chromatin templates that are repressed for AFP transcription in the presence of normal, adult liver extracts or purified, trans-acting factors; and, b) initiating in vitro, remodeling of the repressed AFP gene templates by presentation of cellular extracts or purified trans-activators from AFP expressing sources during semi-conservative DNA replication. c) Activation of chromatin-assembled templates will be determined by subsequent in vitro transcription analysis.
Specific Aim 3 : To employ identified DNA target elements and model assay systems in order to initiate isolation and identification of the protein factors present in hepatic cells and hepatoma cell cultures that confer the dramatic switch in regulated AFP gene expression between normal adult and tumorigenic liver cells. These studies will test the hypotheses that 1) an identified developmental repressor domain is a likely target for inappropriate activation during tumorigenesis and 2) that DNA replication plays an active role in remodeling DNA-protein interactions with dramatic repercussions for AFP gene expression. The long-term goal of our studies is to understand the initiation and progression of tumorigenesis at a molecular level. In pursuing this goal, we will develop an in vitro model of tumorigenic activation of the mouse AFP gene that reconstitutes many properties of the nuclear environment and regulation while remaining biochemically accessible.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM053683-03
Application #
6019122
Study Section
Molecular Biology Study Section (MBY)
Project Start
1997-08-01
Project End
2002-07-31
Budget Start
1999-08-01
Budget End
2000-07-31
Support Year
3
Fiscal Year
1999
Total Cost
Indirect Cost
Name
University of Cincinnati
Department
Genetics
Type
Schools of Medicine
DUNS #
City
Cincinnati
State
OH
Country
United States
Zip Code
45221
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Xie, Zhifang; Zhang, Hai; Tsai, Wenwei et al. (2008) Zinc finger protein ZBTB20 is a key repressor of alpha-fetoprotein gene transcription in liver. Proc Natl Acad Sci U S A 105:10859-64
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Johnson, Amber Buescher; Barton, Michelle Craig (2007) Hypoxia-induced and stress-specific changes in chromatin structure and function. Mutat Res 618:149-62
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Barton, Michelle Craig; Akli, Said; Keyomarsi, Khandan (2006) Deregulation of cyclin E meets dysfunction in p53: closing the escape hatch on breast cancer. J Cell Physiol 209:686-94
Cui, Rutao; Nguyen, Thi T; Taube, Joseph H et al. (2005) Family members p53 and p73 act together in chromatin modification and direct repression of alpha-fetoprotein transcription. J Biol Chem 280:39152-60

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