Dr. William CrosbyDepartment of Biological Sciences, University of Windsor
Systematic profiling of SCF E3 ligase regulatory attributes at play in cell cycle control and cancer
Co-Investigators and Collaborators:
- Dr. Panayiotis Vacratsis, Department of Chemistry and Biochemistry, University of Windsor
- Dr. Wade Harper, Department of Cell Biology, Harvard Medical School
- Dr. Frank Sicheri, Lunenfeld-Tanenbaum, Research Institute, University of Toronto
- Dr. Brian Raught, Ontario Cancer Institute; University of Toronto
EVIDENCE OF PROGRESS
Abnormal presence or absence of specific proteins at different times in the life of a cell can contribute to development of cancer. In many cases, this is caused by the improper regulation of components of those complexes involved in the normal destruction of proteins. This proposal aims to understand how these structures are regulated, assembled, and function.
When proteins are no longer needed or improperly folded in the cell, certain modifications, through a series of enzymes, facilitate their targeted destruction via specific complexes. Defects in genes encoding these enzymes or complexes are implicated in the development of several cancers as well as other diseases. The current state of knowledge in protein degradation biology indicates that there is much to be learned about the regulation of the machinery governing these processes. In collaboration with other labs from Harvard, Toronto, and Windsor, we have recently discovered that specific modifications of a key adaptor protein, Skp1, can act as a switch in regulating the activity of enzymes called E3 ligases. These seemingly independent modifications favour Skp1 interaction with a specific E3 ligase called Cul7 which has been previously shown to be is highly expressed or mutated in lung (81%), breast (70%), skin (59%), ovary (47%), large intestine (44%), and other cancer types.
Our data showed the presence of two novel modification sites on Skp1 protein which are located at the binding interface of Skp1 with other components of E3 ligases. Therefore, the next logical step was to identify the role of these modifications in assembly and activity of the E3 ligase containing Cul7 (CRL7). To address this problem, we proposed three main objectives. First, we identified the precise mechanisms inducing these modifications and determined whether these modifications induced direct interaction of Skp1 with CRL7. Next, we elucidated what other target proteins were recruited to the complex as a result of this interplay. Last, we are addressing the biological effects of the Skp1 and Cul7 interaction by measuring the function of the complex in inducing/inhibiting cancerous characteristics in cells.
Contrary to old assumptions, the Skp1 protein is not just an adaptor protein, but also has regulatory functions in assembly and activity of key complexes involved in critical biological processes such as cellular transformation. We expect this study may lead to new therapeutic approaches for the treatment of cancers with elevated levels of the components of CRL7.
MEASURES OF PROGRESS
A) Manuscripts: Currently working on two manuscripts focusing on the regulatory role of Skp1 on assembly and activity of CRL7.
B) Grants: Will submit to Canadian Cancer Society and to the Cancer Research Society using published data from our study.
C) Training of Highly Qualified Personnel: One post-doctoral fellow was funded through our S4H award.
D) Other Measures: We had the opportunity to work with four distinguished scientists and to get training in the laboratory of one of our collaborators. Our Seeds4Hope research also opened up a new research focus: how E3 ligases are regulated at the transcriptional (i.e., DNA to RNA) level.