- Biological sciences,
Previous work from our laboratory indicates that the Ets family of transcription factors are targets of the ras signal transduction pathway and cooperate with oncogenic ras in cellular transformation. In particular, Ets-2 contains a conserved Map kinase site that becomes phosphorylated in response to the ras/Map kinase pathway. Ets-2phosphorylation leads to the persistent expression of ras-responsive genes. The phosphorylation site resides within a conserved 100 amino acid region referred to as the "pointed" domain. Understanding how phosphorylation leads to Ets-2 activation and the persistent expression of Ets-2 target genes provides insight on the biological effect of oncogenic ras.
We utilized the motheaten mouse model to elucidate the role of Ets-2 in response to signaling in an Erk independent system. In the absence of Erk activity, Ets-2 is phosphorylated on position Thr 72 in the mev mice. We have identified a novel kinase pathway, Akt/JNK, which mediates this phosphorylation. In the me-v cells the complex appears to be constitutively active leading to CSF-1 independent phosphorylation of Ets-2. The kinase activity in WT cells is CSF-1 dependent yet the complex is formed independent of a signal.
To understand the role of Ets-2 phosphorylation, we identified proteins that interact with the "pointed" domain of Ets-2 in a phosphorylation dependent manner. The proteins are members of the mammalian SWI/SNF chromatin-remodeling complex and interact specifically with the unphosphorylated form of Ets-2. Brg-1, a component of theSWI/SNF complex, directly forms a phosphorylation-dependent interaction with the Ets-2 "pointed" domain. Ets-2 and Brg-1 appear to form a transcriptional repressor complex that inhibits the expression of Ras-responsive genes inthe absence of Raf/Erk signaling. When Raf is activated Brg-1 no longer interacts with Ets-2 and a superactivation ofthe uPA reporter is seen.
The expression of the unphosphorylated form of Ets-2 (A72) causes transformation of NIH/3T3 cells. A72 appears to form a transcriptional repressor complex with Brg-1 that is able to transform fibroblasts.
The data presented in this thesis not only provides insight on the regulation of Ets-2 via ras but begins to unravelthe issue of specificity within the Ets family of transcription factors that have a Map kinase site within the "pointed" domain.
Available at: http://works.bepress.com/alicia_schaffner/14/