Figure 2
Truncation studies within the unique N-terminus of PDE4D5 used to probe its interaction with RACK1 using a 2-hybrid screen. Plasmids encoding fusions between the DNA-binding domain of LexA and various amino terminal regions of PDE4D5 were tested for their ability to interact with RACK1, expressed as a fusion with the GAL4 activation domain (left column "pGADN-RACK1"). The identical LexA fusions were tested for their ability to interact with the GAL4 activation domain alone (right column "pGADN"). The regions of PDE4D5 included in the various constructs are annotated. The positions of the portions of PDE4D5 used in this study are shown schematically in Figure 1. The interactions between these components were evaluated with the filter beta-galactosidase assay described previously by us [16, 26]. The bottom two patches serve as internal positive (being the interaction between the oncoproteins RASv12 and RAF) and negative (vectors with inserts) controls, respectively.