Skip to main content
. 2019 Oct 24;8:e50471. doi: 10.7554/eLife.50471

Figure 8. Identification of mutant RhoGDI deficient in extraction of active RhoGTPase.

(A) Oocytes microinjected with halo-tagged WT, Δ51–199, E158/9A and E158/9Q GDI mutants. Scale bar 10 μm, time min:sec; (B) Quantification of GDI intensity at wounds (n = 7–13). Unpaired student’s t-test, 2-tailed distribution, unequal variance statistical analysis to WT. ****p<0.0001. (C) Comparison of Koff values obtained for inactive (Cdc42:GDP, Rho:GDP) and constitutively-active (Cdc42G12V: GTPγS, RhoG14V: GTPγS) Cdc42 and Rho from wash off experiments in presence of either WT (black) or E158/9Q (QQ) (red) GDI. Extraction rates were fitted with a hyperbolic function.

Figure 8—source data 1. Identification of mutant RhoGDI deficient in extraction of active RhoGTPase.

Figure 8.

Figure 8—figure supplement 1. Analysis of previously-described extraction-deficient RhoGDI mutants.

Figure 8—figure supplement 1.

(A–I) Oocytes microinjected with WT or mutant GDI and quantification of localization relative to WT GDI; Unpaired student’s t-test, 2-tailed distribution, unequal variance statistical analysis; (J) Average Koff values obtained for inactive Rho and Cdc42 in absence (control) and presence of 5 μM of either bovine GDI WT or bovine GDI mutants.
Figure 8—figure supplement 1—source data 1. Analysis of previously-described extraction-deficient RhoGDI mutants.
Figure 8—figure supplement 2. Mutant E158/9Q RhoGDI is deficient in extraction of constitutively-active Cdc42Q61L and RhoQ63L in vitro..

Figure 8—figure supplement 2.

(A–B) Comparison of Koff values obtained for inactive (Cdc42:GDP, Rho:GDP) and constitutively-active (Cdc42Q61L:GTP, RhoQ63L:GTP) Cdc42 and Rho from wash off experiments in presence of either WT (black) or E158/9Q (QQ) (red) GDI. Extraction rates were fitted with a hyperbolic function.
Figure 8—figure supplement 2—source data 1. Mutant E158/9Q RhoGDI is deficient in extraction of constitutively-active Cdc42Q61L and RhoQ63Lin vitro.
Figure 8—figure supplement 3. Mutant E163/4Q bovine RhoGDI is deficient in extraction of active Cdc42 and Rho in vitro.

Figure 8—figure supplement 3.

Comparison of Koff values obtained for inactive (Cdc42:GDP, Rho:GDP) and constitutively-active (Cdc42Q61L:GTP, RhoQ63L:GTP) Cdc42 and Rho from wash off experiments in presence of either WT (black) or E163/4Q (QQ) (red) bovine GDI. Extraction rates were fitted with a hyperbolic function.
Figure 8—figure supplement 3—source data 1. Mutant E163/4Q bovine RhoGDI is deficient in extraction of active Cdc42 and Rhoin vitro.