LRRK2 Transport Is Regulated by Its Novel Interacting Partner Rab32

Abstract

Leucine-rich repeat kinase 2 (LRRK2) is a multi-domain 280 kDa protein that is linked to Parkinson's disease (PD). Mutations especially in the GTPase and kinase domains of LRRK2 are the most common causes of heritable PD and are also found in sporadic forms of PD. Although the cellular function of LRRK2 is largely unknown there is increasing evidence that these mutations cause cell death due to autophagic dysfunction and mitochondrial damage. Here, we demonstrate a novel mechanism of LRRK2 binding and transport, which involves the small GTPases Rab32 and Rab38. Rab32 and its closest homologue Rab38 are known to organize the trans-Golgi network and transport of key enzymes in melanogenesis, whereas their function in non-melanogenic cells is still not well understood. Cellular processes such as autophagy, mitochondrial dynamics, phagocytosis or inflammatory processes in the brain have previously been linked to Rab32. Here, we demonstrate that Rab32 and Rab38, but no other GTPase tested, directly interact with LRRK2. GFP-Trap analyses confirmed the interaction of Rab32 with the endogenous LRRK2. In yeast two-hybrid experiments we identified a predicted coiled-coil motif containing region within the aminoterminus of LRRK2 as the possible interacting domain. Fluorescence microscopy demonstrated a co-localization of Rab32 and LRRK2 at recycling endosomes and transport vesicles, while overexpression of a constitutively active mutant of Rab32 led to an increased co-localization with Rab7/9 positive perinuclear late endosomes/MVBs. Subcellular fractionation experiments supported the novel role of Rab32 in LRRK2 late endosomal transport and sorting in the cell. Thus, Rab32 may regulate the physiological functions of LRRK2.

LRRK2 at the interface of autophagosomes, endosomes and lysosomes

“…Rab5 was first found to interact with LRRK2 using a yeast-two-hybrid screening approach [71]. Conversely, LRRK2 was identified as an interaction partner in a yeast-two-hybrid screen for Rab32 [72]. High-throughput protein-protein interaction arrays have shown that LRRK2 physically interacts with Rab7L1 (also known as Rab29) [73].…”

“…In addition, LRRK2 was shown to interact with a number of other Rab GTPases, including Rab32 and Rab38 [72]. Recently, phosphoproteomic screens were performed in an effort to identify bona fide LRRK2 kinase substrates [75].…”

Membrane Trafficking Illuminates a Path to Parkinson’s Disease

“…Further studies using cellular and animal models have suggested that the mutant forms of LRRK2 decrease LC3 lipidation and result in the accumulation of autophagic vacuoles (Roosen and Cookson 2016). In addition, LRRK2 was shown to interact with a number of Rab GTPases, including Rab7, Rab32 and Rab38 (Waschbusch et al 2014). Recently, three functionally related genes (DNAJC13, DNAJC6, and GAK) encoding proteins in the evolutionarily conserved Dnaj/Hsp40 protein family have been identified as the PD-related genes (Cardona and Perez-Tur 2016).…”

Making sense of the cause of Crohn’s – a new look at an old disease

“…It is found in immune cells, in lamina propria macrophages, B-lymphocytes and dendritic cells, and levels are markedly increased in the bowel in CD 320 and in microglia in the nervous system 344 . It interacts with the small GTPases Rab32 and Rab38 with which it co-locates to transport vesicles and recycling endosomes 345 and it is important for the elimination or intracellular Salmonellae 346 and Legionella 347 . Rab32 and Rab38 play an important role in the biogenesis and traffic of melanosomes and lysosomes and this system is disordered in Hermansky-Pudlak syndrome 348 , accounting for the characteristic partial albinism.…”