Figure 6.

The NRD region of c-Myb harbours a SRAF that can switch from being repressive to be activating. (A) CV-1 cells were transfected with 0.2 or 0.4 µg of plasmids expressing Gal4p-DBD (GBD) fused to c-Myb NRD (amino acid residues 410–640) wild-type, 2KR, WT-SUMO-1, 2KR-SUMO-1 or 2EA. The reporter output from the E1b-driven Gal4p-responsive reporter plasmid (5×GRE, 0.2 µg) was normalized to the effect of Gal4p-DBD (0.2 or 0.4 µg), which was set to 100. The results are presented as RLU ± SEM. (B) Based on parallel transfections, using an 1×GRE-E1b-Luc reporter plasmid (0.2 µg), the SFs of the constructs assayed in (A) were calculated. The results are presented as SF ± SEM. (C) CV-1 cells were transfected with plasmids expressing GBD, GBD-NRD or GBD-NRD-2KR together with the SUMO-protease SENP1 or a protease-dead SENP1 mutant. The results are presented as fold change ± SEM, relative to the activity levels of the GBD constructs in the absence of the protease.