Role of autophagy in G2019S‐LRRK2‐associated neurite shortening in differentiated SH‐SY5Y cells

Abstract

Neuritic retraction represents a prominent feature of the degenerative phenotype associated with mutations in leucine rich repeat kinase 2 (LRRK2) that are implicated in autosomal dominant and some cases of sporadic Parkinson’s disease. Alterations in macroautophagy, the vacuolar catabolism of cytoplasmic constituents, have been described in Parkinson’s disease. In this study, we utilized retinoic‐acid differentiated SH‐SY5Y cells to determine whether autophagy contributes to mutant LRRK2‐associated neurite degeneration. Transfection of pre‐differentiated SH‐SY5Y cells with LRRK2 cDNA containing the common G2019S mutation resulted in significant decreases in neurite length, which were not observed in cells transfected with wild type LRRK2 or its kinase‐dead K1906M mutation. G2019S LRRK2 transfected cells also exhibited striking increases in autophagic vacuoles in both neuritic and somatic compartments, as demonstrated by fluorescence and western blot analysis of the autophagy marker green fluorescent protein‐tagged microtubule‐associated protein Light Chain 3 and by transmission electron microscopy. RNA interference knockdown of LC3 or Atg7, two essential components of the conserved autophagy machinery, reversed the effects of G2019S LRRK2 expression on neuronal process length, whereas rapamycin potentiated these effects. The mitogen activated protein kinase/extracellular signal regulated protein kinase (MAPK/ERK) kinase (MEK) inhibitor 1,4‐diamino‐2,3‐dicyano‐1,4‐bis[2‐aminophenylthio]butadiene (U0126) reduced LRRK2‐induced neuritic autophagy and neurite shortening, implicating MAPK/ERK‐related signaling. These results indicate an active role for autophagy in neurite remodeling induced by pathogenic mutation of LRRK2.

Rac1 Protein Rescues Neurite Retraction Caused by G2019S Leucine-rich Repeat Kinase 2 (LRRK2)

“…These studies also suggest that neurite pathology precedes dopaminergic neuronal death by apoptosis (14,15). Although autophagy and increased Tau inclusions have been described in conjunction with neurite shortening, the only signaling pathway linked to the G2019S LRRK2-induced neurite pathology is the ERK pathway (10,15,(17)(18)(19)(20).…”

“…Activated Rac and Cdc42 also signal through PAK to activate LIM kinase, which inhibits downstream cofilin, thereby blocking actin depolymerization (30). Plowey et al (15) found that an ERK inhibitor abolished the G2019S LRRK2-induced neurite retraction in their SH-SY5Y cellular model. PAK signaling through ERK is an interesting upstream trigger that might implicate Rho GTPases in the pathogenesis of neurite shortening.…”

The role of protein clearance mechanisms in organismal ageing and age-related diseases

“…Consequently, the irreversible binding of mutant a-synuclein to the lysosome blocks the binding of other substrates, resulting in a general impairment of the CMA pathway and further a-synuclein accumulation 110 . Proteins related to familiar PD, such as DJ-1 or LRRK2, significantly alter autophagy 111,112 .…”

Autophagy and neurodegeneration

“…In contrast, overexpression of both wild-type LRRK2 and LRRK2 G2019S , but not catalytically inactive forms of the protein, induces an AMPK-mediated upregulation of autophagy (36). Other studies have reported autophagy upregulation following overexpression of LRRK2 G2019S by an ERK1/2-mediated mechanism (37,38).…”