Endogenous Rab38 regulates LRRK2's membrane recruitment and substrate Rab phosphorylation in melanocytes.

Point mutations in leucine-rich repeat kinase 2 (LRRK2) cause Parkinson's disease and augment LRRK2's kinase activity. However, cellular pathways that endogenously enhance LRRK2 kinase function have not been identified. While overexpressed Rab29 draws LRRK2 to Golgi membranes to increase LRRK2 kinase activity, there is little evidence that endogenous Rab29 performs this function under physiological conditions. Here, we identify Rab38 as a novel physiologic regulator of LRRK2 in melanocytes. In mouse melanocytes, which express high levels of Rab38, Rab32, and Rab29, knockdown (or CRISPR knockout) of Rab38, but not Rab32 or Rab29, decreases phosphorylation of multiple LRRK2 substrates, including Rab10 and Rab12, by both endogenous LRRK2 and exogenous Parkinson's disease-mutant LRRK2. In B16-F10 mouse melanoma cells, Rab38 drives LRRK2 membrane association and overexpressed kinase-active LRRK2 shows striking pericentriolar recruitment, which is dependent on the presence of endogenous Rab38 but not Rab32 or Rab29. Consistently, knockdown or mutation of BLOC-3, the guanine nucleotide exchange factor for Rab38 and Rab32, inhibits Rab38's regulation of LRRK2. Deletion or mutation of LRRK2's Rab38-binding site in the N-terminal armadillo domain decreases LRRK2 membrane association, pericentriolar recruitment, and ability to phosphorylate Rab10. In sum, our data identify Rab38 as a physiologic regulator of LRRK2 function and lend support to a model in which LRRK2 plays a central role in Rab GTPase coordination of vesicular trafficking.

The E3 ligase TRIM1 ubiquitinates LRRK2 and controls its localization, degradation, and toxicity.

Missense mutations in leucine-rich repeat kinase 2 (LRRK2) are the most common cause of familial Parkinson's disease (PD); however, pathways regulating LRRK2 subcellular localization, function, and turnover are not fully defined. We performed quantitative mass spectrometry-based interactome studies to identify 48 novel LRRK2 interactors, including the microtubule-associated E3 ubiquitin ligase TRIM1 (tripartite motif family 1). TRIM1 recruits LRRK2 to the microtubule cytoskeleton for ubiquitination and proteasomal degradation by binding LRRK2911-919, a nine amino acid segment within a flexible interdomain region (LRRK2853-981), which we designate the "regulatory loop" (RL). Phosphorylation of LRRK2 Ser910/Ser935 within LRRK2 RL influences LRRK2's association with cytoplasmic 14-3-3 versus microtubule-bound TRIM1. Association with TRIM1 modulates LRRK2's interaction with Rab29 and prevents upregulation of LRRK2 kinase activity by Rab29 in an E3-ligase-dependent manner. Finally, TRIM1 rescues neurite outgrowth deficits caused by PD-driving mutant LRRK2 G2019S. Our data suggest that TRIM1 is a critical regulator of LRRK2, controlling its degradation, localization, binding partners, kinase activity, and cytotoxicity.