of Work: Our group is interested in understanding the role of the Intersectin adaptor protein in regulating signal transduction cascades. Intersectin is an adaptor protein which consists of two NH4-terminal Eps homology (EH) domains, a coil-coil region and 5 tandem Src homology 3 (SH3) domains. Using stable cell lines which express epitope tagged versions of Intersectin, we have shown that Intersectin co-localizes with clathrin suggesting a potential role for Intersectin in endocytosis. Indeed, in collaboration with Brian Kay's laboratory at the University of Wisconsin and Peter McPherson's laboratory at McGill University, we hae shown that overexpression of Intersectin inhibits endocytosis of the transferrin receptor. Furthermore, expression of the SH3 domains also inhibits endocytosis suggesting that this inhibitory activity lies in the SH3 region. Interestingly, the SH3 domains of Intersectin bind dynamin and synaptojanin, two proteins which also play a role in endocytosis. These results suggest that Intersectin may function as a scaffold to assemble endocytotic complexes at clathrin coated pits. Using transient transcriptional reporter assays, we have also shown that Intersectin overexpression leads to activation of the Elk-1 transcription factor. Although Elk-1 is a major transcriptional targt of the MAPK pathway, we have demonstrated through the use of pharmacological inhibitors that Intersectin activates Elk-1 in a MAPK-independent manner. Our current focus is on understanding the mechanism of Intersectin activation of Elk-1 and the importance of this activity for Intersectin function. In addition, given its role in regulation of endocytosis, we are currently examing the possible role of Intersectin in RTK function through regulation of RTK endocytosis, recycling and degradation.

Agency
National Institute of Health (NIH)
Institute
National Institute of Environmental Health Sciences (NIEHS)
Type
Intramural Research (Z01)
Project #
1Z01ES090091-01
Application #
6106797
Study Section
Special Emphasis Panel (LST)
Project Start
Project End
Budget Start
Budget End
Support Year
1
Fiscal Year
1998
Total Cost
Indirect Cost
City
State
Country
United States
Zip Code
Miller, Stephanie L H; Malotky, Erica; O'Bryan, John P (2004) Analysis of the role of ubiquitin-interacting motifs in ubiquitin binding and ubiquitylation. J Biol Chem 279:33528-37
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Rorie, Checo J; Thomas, Venetia D; Chen, Pengchin et al. (2004) The Ews/Fli-1 fusion gene switches the differentiation program of neuroblastomas to Ewing sarcoma/peripheral primitive neuroectodermal tumors. Cancer Res 64:1266-77
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Storey, Nina M; O'Bryan, John P; Armstrong, David L (2002) Rac and Rho mediate opposing hormonal regulation of the ether-a-go-go-related potassium channel. Curr Biol 12:27-33
Oldham, Carla E; Mohney, Robert P; Miller, Stephanie L H et al. (2002) The ubiquitin-interacting motifs target the endocytic adaptor protein epsin for ubiquitination. Curr Biol 12:1112-6
O'Bryan, J P (2001) Determining involvement of Shc proteins in signaling pathways. Methods Enzymol 333:3-15
O'Bryan, J P; Mohney, R P; Oldham, C E (2001) Mitogenesis and endocytosis: What's at the INTERSECTIoN? Oncogene 20:6300-8
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Adams, A; Thorn, J M; Yamabhai, M et al. (2000) Intersectin, an adaptor protein involved in clathrin-mediated endocytosis, activates mitogenic signaling pathways. J Biol Chem 275:27414-20

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