Compromized geranylgeranylation of RhoA and Rac1 in mevalonate kinase deficiency - PubMed (original) (raw)

Compromized geranylgeranylation of RhoA and Rac1 in mevalonate kinase deficiency

L Henneman et al. J Inherit Metab Dis. 2010 Oct.

Abstract

Mevalonate kinase deficiency (MKD) is an autoinflammatory disorder caused by mutations in the MVK gene resulting in decreased activity of the enzyme mevalonate kinase (MK). Although MK is required for biosynthesis of all isoprenoids, in MKD, in particular, the timely synthesis of geranylgeranyl pyrophosphate appears to be compromised. Because small guanosine triphosphatases (GTPases) depend on geranylgeranylation for their proper signaling function, we studied the effect of MK deficiency on geranylgeranylation and activation of the two small GTPases, RhoA and Rac1. We demonstrate that both geranylgeranylation and activation of the two GTPases are more easily disturbed in MKD cells than in control cells when the flux though the isoprenoid biosynthesis pathway is suppressed by low concentrations of simvastatin. The limited capacity of geranylgeranylation in MKD cells readily leads to markedly increased levels of nonisoprenylated and activated GTPases, which will affect proper signaling by these GTPases.

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Figures

Fig. 1

Fig. 1

Effect of simvastatin on the localization and activation of RhoA (a–e) and Rac1 (f–j). Control and mevalonate kinase deficient (MKD) cells were incubated with 0 (white bar; set as 100%), 0.02 (grey bar), or 0.2 μM (black bar) simvastatin for 2 days. Equal amounts of cell lysate (a, f) were analyzed for relative levels of RhoA (b) and Rac1 (g) associated with the membranes and relative soluble levels of RhoA (c) and Rac1 (h). In separate experiments, equal amounts of cell lysate (d, i) were analyzed for relative levels of active RhoA (e) and Rac1 (j). Bars show the mean and standard error of the mean (SEM) of three independent experiments. Immunoblots show the results of one representative experiment. Statistic analysis of observed effects of treatments was performed with one-way analysis of variance followed by Dunnett’s post hoc test. * = P < 0.05, ** = P < 0.01

Fig. 2

Fig. 2

Effect of geranylgeranyltransferase inhibitor (GGTI) on the localization and activation of RhoA (a–e) and Rac1 (f–j). Control and mevalonate kinase deficient (MKD) cells were incubated with 0 μM (white bar; set as 100%) or 20 μM (black bar) GGTI for 2 days. Equal amounts of cell lysate (a, f) were analyzed for relative levels of RhoA (b) and Rac1 (g) associated with membranes and relative soluble levels of RhoA (c) and Rac1 (h). In separate experiments, equal amounts of cell lysate (d, i) were analyzed for relative levels of active RhoA (e) and Rac1 (j). Bars show the mean and standard error of the mean (SEM) of three independent experiments. Immunoblots show the results of one representative experiment. Statistic analysis of observed effects of treatments was performed with paired Student’s t test; * = P < 0.05, ** = P < 0.01

Fig. 3

Fig. 3

Effect of simvastatin on activation of membrane-bound and soluble RhoA (a–c) and Rac1 (d–f). Control and mevalonate kinase deficient (MKD) cells were incubated with 0 (white bar), 0.02 (grey bar), or 0.2 μM (black bar) simvastatin for 2 days. Equal amounts of cell lysate (a, d) were analyzed for relative levels of active RhoA (b) and active Rac1 (e) associated with the membranes (white bar set as 100%) and relative soluble levels of active RhoA (c) and active Rac1 (f) (black bar set as 100%). Bars show the mean and standard error of the mean (SEM) of three independent experiments. Immunoblots show the results of one representative experiment. nd not detected. Statistical analysis of observed treatments effects was performed with one-way analysis of variance followed by Dunnett’s post hoc test; * = P < 0.05, ** = P < 0.01

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