Activin and TGFbeta, members of the TGFbeta superfamily, are pluripotent cytokines that are expressed in virtually every cell of the body. These factors play diverse roles in the body such as regulating early development of the embryo, differentiation, extracellular matrix formation, hematopoiesis, angiogenesis and immune functions. TGFbeta superfamily signaling is transduced by heteromeric serine/threonine kinase receptors at the cell surface and the intracellular mediator, the Smad complex. Following activation of the receptors, there is recruitment and phosphorylation the Smads. As a result the Smad proteins accumulate in the nucleus, bind co-activators or repressors and elicit or suppress transcription of target genes. / To date, the molecular signaling mechanisms for activin/TGFbeta in mammary gland growth and differentiation have not been fully elucidated. Our data identify a novel regulatory crosstalk mechanism by which activin/TGFbeta induced Smad signaling acts to antagonize Stat5 transactivation in mammary epithelial cells. We demonstrate an inhibitory effect of activin/TGFbeta on milk protein expression, specifically betacasein. We further show that activin/TGFbeta inhibitory effect upon betacasein expression is not due to changes in either Stat5 phosphorylation, translocation to the nucleus or binding on the Stat5 response element. We finally demonstrate that the Smads are required to block Stat5 transactivation by activin/TGFbeta and show that they are important mediators in activin/TGFbeta inhibitory response upon Stat5 target gene expression, in particular betacasein and cyclin D1. Finally, we unveil the mechanism by which these two signaling cascades antagonize their effects and find that activated Smads inhibit Stat5 association with its co-activator CBP, thus blocking Stat5 transactivation of its target genes. Thus, we define a novel crosstalk mechanism between two divergent signaling pathways that are involved in regulating mammary gland growth and differentiation. / Whereas the role of TGFbeta signaling in breast cancer has been well characterized, we sought out to study the role and mechanism of action of activin in the human breast cancer T47D cells. We found that activin treatment of T47D cells leads to a potent inhibition of cell growth. We further show that activin induces the Smad, the p38-mitogen activated kinase pathways and the p38 downstream target ATF2. Finally, using specific inhibitors to block p38 MAPK, activin-mediated cell growth inhibition is completely abolished. Together, these results define a novel signaling mechanism induced by activin in breast cancer cells. / Finally in an attempt to identify genes regulated by activin in breast cancer cells, we discover the death adaptor molecule RAIDD as a novel target of activin signaling. We show that RAIDD mRNA and protein levels are potently upregulated by activin. Using antisense-oligos directed against RAIDD, we show that RAIDD expression is necessary in mediating activin inhibition in breast cancer cells. Hence, we define the involvement of a new player in activin mediated cell growth inhibition. / Collectively, these studies reveal novel mechanisms of the activin/TGFbeta signaling cascade in normal mammary epithelial cells and breast cancer cells.
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:QMM.103038 |
| Date | January 2007 |
| Creators | Cocolakis, Eftihia. |
| Publisher | McGill University |
| Source Sets | Library and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada |
| Language | English |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Format | application/pdf |
| Coverage | Doctor of Philosophy (Division of Experimental Medicine.) |
| Rights | © Eftihia Cocolakis, 2007 |
| Relation | alephsysno: 002584313, proquestno: AAINR32343, Theses scanned by UMI/ProQuest. |
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