Ras Project File
Ras is the first oncogene found � a gene involved with cancer � in most human cancers and other animal cancers. Normally, it is active in cellular proliferation and terminal differentiation and is found in most mammals and yeast through a high degree of conservation throughout eukaryotic evolution. There are 3 mutant types: H-ras, for Harvey rat sarcoma; K-ras, for Kirsten rat sarcoma and N-ras. These are present on chromosomes 11, 12 and 1 respectively. Also, these mutant types contain mutations at specific areas that are sometimes notated, with V12, D12, etc. Its protein products are known as p21 proteins are mainly localized in the inner side of the plasma membrane. Its functions include G-binding proteins, conducting GTPase activity and helping signal transduction across the cellular membrane. Ras might potentially be part of a gene superfamily. Ras is activated by autocrine growth signaling, whereby a signal receptor and a signal molecule initiates the Ras pathway, or by a mutation that leaves the Ras in a constant activated state. In autocrine growth signaling, the number of receptor-molecule combinations can be increased to simulate a constant activated state just as mutations would. Mutations to an oncogenic state are induced by retroviruses that lack oncogenic sequences. This disrupts the normal regulatory elements of nearby proto-oncogenes. Secondary mutations may be evolutionary safeguards against excessive transforming properties.
The cancerous state of the Ras gene and its proteins have been combated by using radiation, which sensitizes tumors and turns off the Ras oncogenic pathway (interestingly enough, radiation doesn�t always work), and by using FTIs. Ras must be localized to the plasma membrane as a requirement for their biological activity; this is done by binding of CAAX (C is cysteine, A is an aliphatic amino acid, X is serine or leucine) to FPTase, or farnesyl protein transferase. This is known as farnesylation. This, however, can be blocked by farnesyl transferase inhibitors, or FTIs. Inhibition of this pathway leads to anchorage-dependent and �independent growth of 70% of human tumor cell lines with wild-type and mutant Ras. Certain cell lines are resistant to FTIs, including HeLa, SkBr3 and PANC-1. Testing has shown that sensitive and resistant cell lines are equally susceptible to inhibition of FPTase by FTIs, and that FTIs are capable of suppressing transformation of cancer cell lines with mutations in multiple suppressor and dominant proto-oncogenes, reverting transformed phenotypes back, and being inactive in cells transformed by genes whose products act downstream of Ras. This shows that Ras is effective in some manner and is the gene/protein responsible for the action. Furthermore, there is a correlation between the sensitivity to FTIs of growth-factor mediated signal transduction and cell growth, which could be an argument for autocrine growth signaling as causative of the oncogenic state rather than the mutations (Sepp-Lorenzino, Cancer Research 55:5302-9).
Project Goal
FTIs have been shown to work with mutated Ras that has, by necessity, conjured up an infinite activated state that causes unlimited growth of the cancer. It also works with wild-type Ras. It has never been shown if these are activated as well. The goal for this project is to determine if there is an activation of wild-type Ras when using FTIs, and so take a step in concluding if FTIs work by a Ras-dependent or �independent means. There is a problem which will have to be combated; other proteins also rely on farnesylation and not just Ras, so the anti-cancer properties of FTI might be caused by interaction with another protein.