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Dissertation
Regulation of tyrosine hydroxylase gene expression by aFGF and co-activators in the developing neurons
Doctor of Philosophy (Ph.D.), Allegheny University of the Health Sciences
May 1997
DOI:
https://doi.org/10.17918/00008978
Abstract
Previous studies from this laboratory have demonstrated that the catecholamine (CA) biosynthetic Enzyme tyrosine hydroxylase (TH) can be induced in non-CA neurons of the striatum by the synergistic interaction of acidic fibroblast growth factor (aFGF) and a number of different co-activators (dopamine, TPA, IBMX/forskolin). In my study, I sought to determine the underlying molecular mechanisms for this novel expression by identifying the relevant cis-acting DNA sequences of TH gene and transcription factors. To do so, cultures of E14 rat striatum were grown one day in culture in defined medium prior to treatment with aFGF (100ng/ml) and/or dopamine (20 uM), TPA (200 nM), IBMX (0.25 mM)/forskolin (50 uM). Sixty minutes after stimulation, cultures were harvested, nuclear extracts were prepared and analyzed by gel shift, supershift assays and Western blotting. We found that the binding of transcription factors to the TH-AP-1, TH-CRE-2, TH-TATA was increased by aFGF and co-activator stimulation as compared to control (media only), while the binding of TH-CRE proteins remained unchanged. To further investigate the components of the TH-AP-1, TH-CRE, TH-CRE-2 protein complexes, supershift assays using antibodies to specific transcription factors were performed. We discovered that in the TH-AP-1 site, the binding of Fos-B and Jun-D was increased, c-Fos and other unknown Fos was recruited to the AP-1 site, decrease in binding of ATF-2 and CREM was also revealed, while the binding of c-Jun remained the same after stimulation by aFGF and co-activators. However, stimulation of the cultures with aFGF and co-activators also resulted in the novel recruitment of c-Fos; the increased binding of ATF-2 and CREM to the TH-CRE site, although the c-Jun and Jun-D existed in the same amount within CRE protein complexes in stimulated striatal neurons as compared to the control. In addition, we also found the increase in binding of CREM to the TH-CRE-2 site following the treatment with aFGF and co-activators, even though the majority of CRE-2 protein complexes is still unknown. Therefore, we concluded that the protein complexes of AP-1, CRE, and CRE-2 interact directly or indirectly with the basal transcription complexes to initiate the novel expression of TH gene in striatal neurons by aFGF and co-activators.
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Details
- Title
- Regulation of tyrosine hydroxylase gene expression by aFGF and co-activators in the developing neurons
- Creators
- Zheng Guo
- Awarding Institution
- Allegheny University of the Health Sciences
- Degree Awarded
- Doctor of Philosophy (Ph.D.)
- Publisher
- Allegheny University of the Health Sciences; Philadelphia, Pennsylvania
- Number of pages
- xiii, 200 unnumbered pages
- Resource Type
- Dissertation
- Language
- English
- Academic Unit
- Anatomy (and Neurobiology) [Historical]; Allegheny University of the Health Sciences (1996-1998); School of Medicine (1996-1998)
- Other Identifier
- 991021888862604721