The TGF-beta family of ligands signal through a unique heteromeric receptor complex distinguished by its serine-threonine kinase activity and a novel family cytoplasmic proteins termed Smads. In this pathway, receptor-activated Smads2 and 3 are phosphorylated directly by the type I receptor kinase and, in association with a common mediator Smad4, translocate to the nucleus where they participate in transcriptional complexes. We have taken a multi-faceted approach to gain insight into the biochemistry of this pathway in vitro and to understand its significance in vivo. One approach has been to identify unique factors that modulate receptor or Smad activity either directly or indirectly. A novel nuclear protein first identified in this laboratory, SNIP1, has been shown to interact with Smad4 and to suppress TGF-beta signaling, and now, based on a yeast two-hybrid screen, to interact with an important TGF-beta target, c-Myc to enhance its transcriptional activating activity. SNIP1 stabilizes c-Myc and enhances its interaction with the coactivator, p300. Ongoing studies examining the expression patterns of SNIP1 in a variety of human cancers suggest that it will play a complex role both in blocking the tumor suppressor functions of TGF-beta and enhancing the oncogenicity of c-Myc. To complement the above basic biochemical approaches, we have also developed a strong program of research based on the hypothesis that deletion of specific downstream signaling pathways in vivo should, conceptually, have a less severe and more selective effect than broader-based approaches involving targeted deletion or overexpression of ligand or receptors. Processes targeted include wound healing, fibrosis, and especially carcinogenesis. In one approach we are attempting to identify specific targets of Smad2 and Smad3 in cancer cells, by characterizing the outcome of selective overexpression or suppression of Smad2 or Smad3 in tumorigenesis and metastasis. Much of this effort is presently focused on examining both in vitro and in vivo effects of altering the balance of these two pathways in human breast cancer cells derived from the parental MCF10A line. Initial results show that the Smad pathway mediates both tumor suppressor and pro-metastatic activities of TGF-beta on tumor cells, though specific roles of Smad2 and Smad3 remain to be identified. Another approach is based on in vitro co-culture of tumor cells and stromal cells, where the genotype of each component cell can be manipulated. Initial results show that MMP-9 is produced by fibroblasts in response to signals from tumor cells and depends both on the degree of tumorigenicity of the epithelial component and the signaling pathways in the fibroblasts. We have shown that both MMP-9 secretion and migration of the tumor cells is dependent, in part, on TGF-beta signaling through Smad3 as well as through MAPK pathways. Another aspect of the stromal response that we are investigating involves understanding mechanisms of suppression of immune surveillance by TGF-beta, especially those involving dendritic cells.The Smad3 knockout mouse, developed by Chuxia Deng, NIDDK, continues to provide new insights into the roles of TGF-beta in repair of injury and in fibrosis. We are using a novel wound model in the ear to unravel novel roles of Smad3 in modulation of mechanotransduction of wound closure dependent on changes in the extracellular matrix, including especially elastin, collagen, glycosaminoglycans and the integrins.

Agency
National Institute of Health (NIH)
Institute
Division of Basic Sciences - NCI (NCI)
Type
Intramural Research (Z01)
Project #
1Z01BC005051-03
Application #
7337751
Study Section
(LCRC)
Project Start
Project End
Budget Start
Budget End
Support Year
3
Fiscal Year
2006
Total Cost
Indirect Cost
Name
Basic Sciences
Department
Type
DUNS #
City
State
Country
United States
Zip Code
Saika, Shizuya; Yamanaka, Osamu; Ikeda, Kazuo et al. (2005) Inhibition of p38MAP kinase suppresses fibrotic reaction of retinal pigment epithelial cells. Lab Invest 85:838-50
Stuelten, Christina H; DaCosta Byfield, Stacey; Arany, Praveen R et al. (2005) Breast cancer cells induce stromal fibroblasts to express MMP-9 via secretion of TNF-alpha and TGF-beta. J Cell Sci 118:2143-53
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