Rabaptin-5 levels are decreased in the absence of RabGEF1. (A) Total cell lysates from RabGEF1 +/+ and −/− BMCMCs were analyzed by Western blot using α–Rabaptin-5, α-RabGEF1, and α-Rab5 Abs (left panel). The blot was reprobed with α-GAPDH Abs to show loading. Results are representative of those obtained in 6 or more experiments. Rabaptin-5 signals from 6 separate experiments were quantified by densitometric scanning and corrected for loading (right panel). (B) Rabaptin-5 mRNA levels in +/+ and −/− BMCMCs were analyzed by semiquantitative RT-PCR (28 cycles; left panel). Signals from 3 separate experiments were quantified by densitometric scanning and corrected for loading (right panel). (C) Total cell lysates from Figure 2 B (RabGEF1 +/+ and −/− BMCMCs as well as −/− BMCMCs expressing the indicated RabGEF1 mutants) were analyzed by Western blot using α–Rabaptin-5 Abs (left panel). The blot was reprobed with α-GAPDH Abs to show loading. Rabaptin-5 signals from 4 (NT and D313A) or 5 separate experiments were quantified by densitometric scanning and corrected for loading (right panel). (A-C; right panels) Data shown are the mean + SEM. (A) ++ indicates P < .01 versus +/+. (C) + indicates P < .05; +++, P < .001 versus +/+ “empty” values; * indicate P < .05; **, P < .01; ***, P < .001 versus−/− “empty” values.