Yeast Two-Hybrid Screening as a Means of Deciphering Tumor Suppressor Pathways
With the identification and functional characterization of various tumor suppressor genes, the multiprotein pathways in which they function are beginning to be delineated. Mutations in critical pathways, such as the basal cell cycle machinery, DNA surveillance and repair, apoptosis, and checkpoint control ultimately provide a selective growth advantage leading to unfettered cell proliferation and tumor formation. Understanding the cross-talk between these pathways as well as protein-protein interactions within the pathways themselves has become increasingly important in order to design better therapeutic strategies. Therefore methods involved in identifying novel protein-protein interactions in mammalian cells have become just as valuable as those that served initially to identify the tumor supresssor genes. Large-scale biochemical purification of interacting proteins is a viable strategy that was, in one case, used to identify a number of novel proteins (i.e., TRAF1, TRAF2, c-IAP1, and c-IAP2) (1 ,2 ) that interact with the tumor necrosis factor receptor 2. However, due to the difficulty of reconstituting in-vivo interactions in a large-scale purification scheme as well as still having to clone your gene of interest, this method can prove to be difficult as well as time-consuming.