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M.S., Fudan University -2005
Thesis Advisor: Joseph D. Fondell, Ph.D.
Graduate Program in Physiology & Integrative Biology
2nd floor Conference Room
Wednesday, June 20, 2012
MED1 is a key component of the Mediator coactivator complex that bridges gene-specific
activators with RNA polymerase II and the basal transcription machinery. In cultured prostate
cancer cells, MED1 targets Mediator to the androgen receptor (AR) and is essential for robust
androgen-dependent transcription. Notably, MED1 is overexpressed in a number of prostate
cancer cell lines and specimens, yet its role in clinical prostate tumorigenesis remains unclear.
Furthermore, the mechanisms by which MED1 contacts AR are obscure and the spectrum of AR
target genes that are specifically coactivated by MED1-Mediator remain poorly defined.
In this study, we set out to investigate the coregulatory role of MED1 in prostate cancer cells.
By performing RNA profiling assays in prostate cancer cells in which MED1 levels were silenced by
RNAi, we identified a number of known and novel AR-target genes that are coactivated by MED1-
Mediator including Cdc6 which encodes a licensing factor for DNA replication. We further
employed chromatin immunoprecipitation (ChIP) and electromobility shift assay (EMSA) to
precisely characterize a critical androgen response element in the Cdc6 promoter (-734 to -720 bp
upstream of the transcription start site). By utilizing a series of AR and MED1 point and deletion
mutants, we investigated how the two proteins precisely bind to one another in the presence of
ligand. Significantly, we show for the first time that MED1 binds to a distinct AR N-terminal domain
region termed transactivation unit-1 (Tau-1) via two newly discovered noncanonical á-helical motifs
located between MED1 residues 505 and 537.
Finally, by staining human prostate cancer tissue microarrays with antibodies against MED1,
we demonstrate that MED1 expression levels are elevated in the malignant epithelium of
statistically significant numbers of prostate cancer patients and that MED1 overexpression
correlates with increased cellular proliferation. We also found that ectopic MED1 overexpression in
human prostate cancer xenografts significantly promotes tumor growth in nude mice. Towards
addressing the underlying mechanism by which MED1 becomes overexpressed in prostate cancer,
we found that suppression of either AKT or ERK kinase activity with specific inhibitors decreased
MED1 expression in situ, whereas forced overexpression of constitutively active ERK or AKT
increased MED1 expression. Moreover, in a Nkx3.1;Pten mutant mouse model of prostate cancer,
in which ERK and AKT signaling pathways are genetically programmed to become hyper-activated
in parallel with disease progression, we found that MED1 levels were nearly double that observed
in wild type prostates. The data is striking in light of earlier studies showing that MED1
phosphorylation not only stabilizes its nuclear half-life, it enhances its coactivator activity as well.
Collectively, this study provides new insights into the mechanisms and activity of AR-MED1-
Mediator complexes that should have clinical implications for AR mediated gene expression in