Investigating Rab8 Phosphorylation and AMPA Receptor Trafficking in the LRRK2G2019S Mutation

PROJECT ABSTRACT Kinase enhancing mutations in Leucine-Rich Repeat Kinase 2 (LRRK2) are associated with both familial and sporadic forms of Parkinson’s (PD), including the prevalent G2019S mutation. In brain, LRRK2 is highly expressed in striatal projection neurons (SPNs) and cortical neurons. Our lab has previously shown significant AMPA receptor (AMPAR) trafficking deficits in mouse Lrrk2G2019S SPNs, however the molecular mechanisms underlying these trafficking deficits is unknown. LRRK2 phosphorylates a number of Rab GTPases, including Rab8, and previous studies in hippocampal neurons suggest a role for Rab8 in AMPAR trafficking. Rab8’s interactions with specialized proteins that maintain a tightly regulated activation cycle are impaired by LRRK2- mediated phosphorylation, suggesting that Rab8 activity is further regulated by LRRK2 phosphorylation. How Rab8 regulates AMPAR trafficking in SPNs, and the functional consequences of LRRK2-mediated phosphorylation of Rab8, are not known. Preliminary evidence suggests that both phosphorylated and unphosphorylated Rab8 interact with the AMPAR subunit GluA1 in vitro. Therefore, I hypothesize that Rab8 phosphorylation state regulates AMPAR trafficking in SPNs, and excess phosphorylated Rab8 (p72-Rab8; pRab8) mediates AMPAR trafficking deficits seen in the Lrrk2G2019S mutation. To address these gaps, the proposed experiments aim to characterize Rab8’s functional relationship with AMPAR trafficking pathways in SPNs and explore the effects of LRRK2-mediated phosphorylation of Rab8 by combining genetic manipulations of Rab8 with imaging, biochemical, and electrophysiological approaches.