Expression of GFP-rab8WT and GFP-rab8Q67L
downregulates the expression of endogenous rab8 in transgenic tadpoles.
Equivalents of 1/8 of the retinas isolated from stage 51 tadpoles were
subjected to SDS-PAGE and immunoblotted with anti-rab8
antibody (left panel) and anti-GFP antibody (right panel). In addition
to a 23-kDa protein recognized by anti-rab8 antibody, transgenic
retinas contain a 50-kDa protein that is also recognized by anti-GFP.
The expression of GFP-rab8WT fusion protein under the control of the
X. laevis rhodopsin promoter does not exceed that of
endogenous rab8 as measured in the nontransgenic retina. However, the
amount of the endogenous rab8 in these tadpoles is <50% of the
nontransgenic animal. Therefore, the total content of rab8 (endogenous
plus the fusion protein) is ∼1.5- to 2-fold higher than in the
nontransgenic animals. In GFP-rab8Q67L tadpoles endogenous rab8 is also
reduced to similar levels (<50% of control) as in GFP-rab8WT animals,
and the fusion protein is in a threefold excess over the endogenous
rab8. Because of the progressive retinal degeneration in GFP-rab8Q67L
animals, the contribution of rod photoreceptors is lower than that in
the retinas expressing GFP-rab8WT protein.