Figure 4.

Decreased Rabggta expression and Rab GGTase activity in gm/gm mice. (A) Western blots of wild-type (+/+), heterozygous (gm/+), and mutant (gm/gm) platelets probed with Rabggta and β-actin antibodies. Rabggta (60 kDa) was reduced ≈70% in gm/gm. This experiment was repeated four times with the same results. (B) Rab GGTase activity of gm/gm and +/+ platelets. Duplicate cytosolic protein extracts from two samples of pooled platelets from gm (open symbols) and C57BL/6J mice (solid symbols) were assayed for Rab GGTase activity by measuring transfer of [³H]geranylgeranyl ([³H]GG) to Rab1a (15). This experiment was repeated four times with platelets, and twice with liver, spleen, and kidney protein extracts with similar results. (C) GGTase-I activity of gm/gm and +/+ pooled platelets. GGTase-I activity was determined by measuring transfer of [³H]GG to Rac1. Each point represents an average of duplicate reactions. For Rab GGTase reactions, a control reaction incubated after addition of 50 mM EDTA was subtracted from all samples. For GGTase-I reactions, a control reaction incubated with buffer alone was subtracted from all samples. These experiments were repeated at least twice in each of the liver, kidney, and spleen extracts with similar results to platelets.