Mapping of Toca-1 binding site in N-WASP.

• Main Author: Liong, Kai Ni.
• Other Authors: Thirumaran s/o Thanabalu
• Format: Final Year Project
• Language: English
• Published: 2013
• Subjects: DRNTU::Science
• Online Access: http://hdl.handle.net/10356/54803
• Institution: Nanyang Technological University

Cancer metastasis contributes greatly to the mortality of cancer patients. It is strongly associated with the formation of actin-rich matrix degrading surface protrusions, called invadopodia, by metastatic carcinoma cells. They resemble podosome produced by immune cells and mediate locomotion through dense extracellular matrix (ECM) by playing a crucial role in ECM remodelling. Under the influence of epidermal growth factor (EGF), invadopodia is formed, with N-WASP and Arp2/3 as the core proteins. Toca-1 binds and regulates the activity of N-WASP. Studies had shown that both Toca-1 and N-WASP were upregulated in invasive tumor cells. Together, they form complex to promote actin polymerization. It is widely known that Toca-1 binds to prolin-rich region of N-WASP through its SH3 domain; however, the exact binding site has not been mapped. In this study, Gal4-Yeast Two Hybrid assay was used to identify the binding site(s) of Toca-1 in N-WASP. It was found that Toca-1 was able to bind to 5 putative sites in N-WASP1-505. They are found within N-WASP8-11, N-WASP309-317, N-WASP317-330, N-WASP356-365 and N-WASP365-384.