The Parkinson’s disease-associated protein, leucine-rich repeat kinase 2 (LRRK2), is an authentic GTPase thatstimulates kinase activity

Abstract

Mutations in the leucine-rich repeat kinase 2 (LRRK2) gene are the leading cause of autosomal dominant Parkinson's disease (PD). LRRK2, a member of the ROCO protein family, contains both Ras GTPase-like (Roc) and kinase (MAPKKK) domains, as well as other functional motifs. Here, we have identified LRRK2 as the first mammalian ROCO protein that is an authentic and functional GTPase, defined by the ability to bind GTP and undergo intrinsic GTP hydrolysis. Furthermore, the Roc domain is sufficient for this native GTPase activity and binds and hydrolyzes GTP indistinguishably from the Ras-related small GTPase, Rac1. The PD-associated mutation, R1441C, located within the Roc domain, leads to an increase in LRRK2 kinase activity and a decrease in the rate of GTP hydrolysis, compared to the wild-type protein, in an in vitro assay. This finding suggests that the R1441C mutation may help stabilize an activated state of LRRK2. Additionally, LRRK2 mediated phosphorylation is stimulated upon binding of non-hydrolyzable GTP analogs, suggesting that LRRK2 is a MAPKKK activated intramolecularly by its own GTPase. Since GTPases and MAPKKKs are upstream regulators of multiple signal transduction cascades, LRRK2 may play a central role in integrating pathways involved in neuronal cell signaling and the pathogenesis of PD.

Ancient Origin of the Parkinson Disease Gene LRRK2

“…In humans, however, it is unknown whether loss-of-function mutations in LRRK2 lead to dopaminergic cell death. On the contrary, it seems that it is increased/constitutive activity of the LRRK2 protein (West et al 2005;Gloeckner et al 2006;Smith et al 2006;Guo et al 2007;Li et al 2007). In summary, researchers in this field should be very careful before extrapolating results obtained for protostome LRRK genes to humans.…”

Is inhibition of kinase activity the only therapeutic strategy for LRRK2-associated Parkinson's disease?

“…The GTPase activity for N1437H has not been determined, but it has been shown that this mutant has increased GTP binding [13], which is consistent with lower GTPase activity. The reported GTP-binding capacity of R1441C/G and Y1699C is less consistent, with some reports of an increase and others finding no change in GTP binding [45,47,48]. These effects are probably related, as it has been proposed that the ROC and COR domains may interact with each other.…”

Update on the Functional Biology of Lrrk2

“…The I2020T mutation has been reported to increase kinase activity [44,57] but also to reduce kinase activity [55]. Data on kinase alterations by mutation at position R1441 are conflicting, with reports of no change [20,55,58] and increased [47,57,59] for R1441C; and unaltered [55] and increased [57] for R1441G. Finally, the Y1699C mutation is reported as unaltered [20,55] and increased [57].…”

“…Ras-GTPases act as molecular switches that cycle between GTP-bound (active) and GDPbound (inactive) conformations, a process that regulates many diverse cell signaling functions. This process is additionally facilitated by guanine nucleotide exchange factors (GEFs) and GTPase-activating proteins (GAPs) [59 ].…”

“…Several groups now have shown that Lrrk2 can bind GTP via GTP-sepharose or GTPγS-binding assays, whereas disruption of the P-loop in the nucleotide-binding pocket of the Roc domain abolishes GTP binding [33,47,53,[57][58][59][60]. GTPase activity has been more difficult to demonstrate [57,60], presumably owing to the requirement of GEFs and GAPs for in vitro assays; however, optimized experimental conditions have now enabled GTPase activity, albeit low, to be detected [33,53,58,59].…”