Molecular Mechanisms Of Estrogen-Dependent Transcriptional Regulation Through Ap-1
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Isaacs, Gary David
Gene regulation by estrogen (E2) plays an important role in mediating physiological responses in normal and disease states. Estrogen receptors (ERs) facilitate these responses by at least two genomic modes of action: (1) binding directly to estrogen response elements (EREs) or (2) binding indirectly to DNA through transcription factors (like AP-1). Although the ERE pathway is well studied, little is known about the mechanism of E2-dependent actions through this indirect pathway (e.g., ER/AP-1 pathway). Using an unbiased proteomic approach, that utilizes affinity purification and iTRAQ labeling, I examine the composition of AP-1 complexes in order to better understand the foundation of ER tethering. The results from this analysis identify several AP-1 and non-AP-1 transcription factors associated with E2-responsive AP-1 sites. My results also identify putative coregulators that may play a role in mediating ER/AP-1 responses in vivo. In further studies, I characterize the genomic interplay between E2-signaling and the AP-1 regulator, Jun N-terminal Kinase 1 (JNK1). Interestingly, I show that JNK1 binds to discrete regions of the genome in an E2-regulated manner and correlate these binding events with ER-alpha occupancy. I also define the transcription factors responsible for tethering JNK1 to promoter regions. These results reveal the emerging theme that MAP kinases (like JNK1) can form stable, chromatin-associated complexes. Furthermore, I describe the necessity of JNK1 activity in mediating E2- dependent transcriptional outcomes in breast cancer cells and demonstrate the importance of JNK1 in E2-dependent tumor cell growth. Finally, I show that JNK1 can phosphorylate coactivators involved in E2-dependent complexes, as well as histone H3. Modification of these factors may play a role in facilitating E2-dependent transcriptional responses in vivo. My results establish a new paradigm for estrogen signaling which now includes JNK1 as an E2-dependent coregulator.
dissertation or thesis