The RAS/RAF/MEK/ERK pathway is the prototypical cellular signal transduction cascade and has been the focus of intense scrutiny over the last two decades. As a mitogenic pathway, its activation is a potent driver of cellular growth and survival, and its deregulation underlies many cancers. While RAS family GTPases have long been recognized as prolific human oncogenes, a landmark study in 2002 also established the RAF family kinase as a bona fide oncogene (Davies et al., 2002). Indeed, aberrant RAS-RAF signaling underlies nearly one-third of all human cancers (Wellbrock et al., 2004). Notably, mutations in RAF are found with astounding frequency in certain cancers (e.g. 70% of malignant melanomas) (Dhomen and Marais, 2007). These findings have identified intercepting aberrant RAF function as an ideal therapeutic target. RAF is a Ser/Thr protein kinase and its activity is strictly regulated by a core complex of at least three proteins, namely, KSR, CNK and HYP (Claperon and Therrien, 2007). The mechanism by which the KSR/CNK/HYP complex regulates RAF function remains enigmatic. In particular, the function of KSR in regulating RAF activity is highly controversial. The work described in this thesis was conducted with the aim of: i) understanding the interactions that underlie formation of the KSR/CNK/HYP complex, and ii) elucidating the mechanism by which the complex regulates RAF function. I have attempted to accomplish these aims using a combination of structural biology, biochemistry and cell biology approaches. I begin by presenting the structure of the SAM domain mediated interaction between CNK and HYP. I describe a model for how the CNK/HYP interaction in turn serves to recruit KSR to form the higher-order KSR/CNK/HYP complex.
Subsequently, I describe the allosteric mechanism by which KSR controls RAF activation via the formation of specific side-to-side kinase domain heterodimers of KSR and RAF. Lastly, I describe a potential mechanism by which RAS directly mediates the attainment of the side-to-side dimer configuration of RAF through its own ability to form dimers. The acquisition of the side-to-side dimer configuration is essential for aberrant RAF signaling in cancers, suggesting future RAF inhibition strategies could be aimed at preventing dimer formation.
| Identifer | oai:union.ndltd.org:TORONTO/oai:tspace.library.utoronto.ca:1807/42552 |
| Date | 19 November 2013 |
| Creators | Rajakulendran, Thanashan |
| Contributors | Sicheri, Frank |
| Source Sets | University of Toronto |
| Language | en_ca |
| Detected Language | English |
| Type | Thesis |
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