Figure 2 of
Tucker, Mol Vis 2004;
Figure 2. Dominant negative behavior is seen in recessive GUCY2D mutations L954P and P858S
Co-expression of the catalytic domain mutants L954P and P858S with wild-type RetGC-1 reveals dominant-negative behavior. HEK293 cells were transfected with 2.5 μg wild-type RetGC-1 and either 2.5 or 5 μg of pRC-CMV, P858S, or L954P. A: Constructs were transiently transfected in HEK 293 cells, and harvested membranes were immunoblotted with an antibody to the RetGC-1 kinase homology domain (Inset). B: RetGC-1 activity was assessed in membranes incubated in GC buffer with 1.7 μM GCAP-2 and 1 mM EGTA (e.g., GCAP-2 stimulation). C: RetGC-1 activity was assessed in membranes incubated in GC buffer with 1% Triton X-100 and 10 mM MnCl2 instead of MgCl2 (e.g., Mn/Triton stimulation).