GTPase Activity Plays a Key Role in the Pathobiology of LRRK2
Figure 5
GTPase activity modulates LRRK2-induced neuronal toxicity.
(A) Human LRRK2 domain fragments containing the GTPase domain (GTP and GTP-COR-Kin) but not the kinase domain (Kin) alone induce neuronal toxicity similar to full-length WT LRRK2. Representative fluorescent images (eGFP) showing mouse primary cortical neurons co-transfected with LRRK2 constructs and eGFP in a 10∶1 molar ratio or transduced with HSV-WT-LRRK2/CMV-eGFP virus expressing full-length WT LRRK2. Neuronal viability was analyzed at 48 hrs post-transfection (DIV 12) with non-viable neurons exhibiting obvious neurite process and/or nuclear fragmentation (arrows). (B) Quantification of neuronal viability induced by LRRK2 expression. Bars indicate the viability of eGFP-positive neurons (n = 200) for each transfection condition expressed as a percent (%) of control neurons (eGFP only). Data represent the mean ± SEM from three independent experiments. Data were analyzed for statistical significance by two-tailed unpaired Student's t-test compared to control neurons (*P<0.01 and **P<0.001). n.s., non-significant. (C) LRRK2 GTPase variants (in GTP-COR-Kin fragment) induce neuronal toxicity. Representative fluorescent images (eGFP) of neurons at 48 hrs post-transfection containing truncated LRRK2 GTPase variants and eGFP. Arrows indicate non-viable neurons. (D) Quantification of neuronal viability induced by truncated LRRK2 GTPase variants. Bars indicate the viability of eGFP–positive neurons (n = 200) for each transfection condition expressed as a percent (%) of control neurons (eGFP only). Data represent the mean ± SEM from three independent experiments. Data were analyzed for statistical significance by two-tailed unpaired Student's t-test compared to control neurons, or by pair-wise comparisons where indicated by horizontal lines (*P<0.05 and **P<0.005). n.s., non-significant versus control.