Abstract
Pathologic inclusions composed of α-synuclein called Lewy pathology are hallmarks of Parkinson’s Disease (PD). Dominant inherited mutations in leucine rich repeat kinase 2 (LRRK2) are the most common genetic cause of PD. Lewy pathology is found in the majority of individuals with LRRK2-PD, particularly those with the G2019S-LRRK2 mutation. Lewy pathology in LRRK2-PD associates with increased non-motor symptoms such as cognitive deficits, anxiety, and orthostatic hypotension. Thus, understanding the relationship between LRRK2 and α-synuclein could be important for determining the mechanisms of non-motor symptoms. In PD models, expression of mutant LRRK2 reduces membrane localization of α-synuclein, and enhances formation of pathologic α-synuclein, particularly when synaptic activity is increased. α-Synuclein and LRRK2 both localize to the presynaptic terminal. LRRK2 plays a role in membrane traffic, including axonal transport, and therefore may influence α-synuclein synaptic localization. This study shows that LRRK2 kinase activity influences α-synuclein targeting to the presynaptic terminal. We used the selective LRRK2 kinase inhibitors, MLi-2 and PF-06685360 (PF-360) to determine the impact of reduced LRRK2 kinase activity on presynaptic localization of α-synuclein. Expansion microscopy (ExM) in primary hippocampal cultures and the mouse striatum, in vivo, was used to more precisely resolve the presynaptic localization of α-synuclein. Live imaging of axonal transport of α-synuclein-GFP was used to investigate the impact of LRRK2 kinase inhibition on α-synuclein axonal transport towards the presynaptic terminal. Reduced LRRK2 kinase activity increases α-synuclein overlap with presynaptic markers in primary neurons, and increases anterograde axonal transport of α-synuclein-GFP. In vivo, LRRK2 inhibition increases α-synuclein overlap with glutamatergic, cortico-striatal terminals, and dopaminergic nigral-striatal presynaptic terminals. The findings suggest that LRRK2 kinase activity plays a role in axonal transport, and presynaptic targeting of α-synuclein. These data provide potential mechanisms by which LRRK2-mediated perturbations of α-synuclein localization could cause pathology in both LRRK2-PD, and idiopathic PD.
Highlights
Inclusions composed primarily of the protein, ɑ-synuclein, found in axons, called Lewy neurites, and in the soma, called Lewy bodies [1], are among the primary hallmarks of Parkinson’s Disease (PD)
leucine rich repeat kinase 2 (LRRK2) kinase inhibition increases colocalization of α‐synuclein and the presynaptic marker Vesicular glutamate transporter 1 (vGLUT1) in primary hippocampal neurons α-Synuclein is highly expressed in glutamatergic presynaptic terminals where it colocalizes with vGLUT1 [15, Table 1 List of statistical tests and results for each figure vGLUT1/ α-synuclein vGLUT1 α-synuclein vGLUT1/ α-synuclein vGLUT1 α-synuclein α-synuclein LP2
Understanding LRRK2 kinase activity could be potentially important for putative treatment options of both LRRK2-PD and sporadic PD cases [4], Given that tetrameric, native α-synuclein is resistant to aggregation [30, 38, 39], our data suggest that LRRK2 kinase inhibitors could have a therapeutic benefit in halting Parkinson’s disease progression and development of nonmotor symptoms by increasing native, presynaptic α-synuclein at membranes and protecting against pathologic aggregation. Both LRRK2 and α-synuclein localize to the presynaptic terminal, and LRRK2 kinase activity has been shown by multiple studies to impact pathologic α-synuclein aggregation, but how LRRK2 kinase influences normal α-synuclein has remained unclear
Summary
Inclusions composed primarily of the protein, ɑ-synuclein, found in axons, called Lewy neurites, and in the soma, called Lewy bodies [1], are among the primary hallmarks of PD. Mutations in LRRK2 cause familial PD, and increase its kinase activity. LRRK2 kinase activity has been observed in post mortem tissue analysis of idiopathic PD brains [4]. Lewy pathology is common in individuals with PD harboring the most common G2019S-LRRK2 mutation, which is associated with increased nonmotor symptoms [5]. PD animal models demonstrate α-synuclein pathologic aggregation is exacerbated by G2019S-LRRK2 expression [6,7,8,9,10,11]. Given the importance of LRRK2 activity in both familial, and idiopathic PD cases, deciphering the downstream effects of LRRK2 kinase activity is crucial for investigating PD mechanisms, in general. An understanding of how LRRK2 kinase activity interacts with α-synuclein could help with determine how these two proteins contribute to LRRK2-PD, and idiopathic PD
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