Objective: To elucidate the molecular basis of the Park20 by evaluating the role of synaptojanin 1 in intracellular trafficking.

Background: Recently a new form of autosomic recessive parkinsonism (PARK20) has been reported due to mutation in SYNJ1. This gene encodes for synaptojanin 1, a phosphoinositide phosphatase, involved in recycling of synaptic vesicles in neurons.

Methods: We assessed the effects of SYNJ1 knockdown on the endocytic pathway evaluating the morphology and function of endosomal compartments through quantitative confocal analysis and internalization assays. We carried out the study in human neural (SH-SY5Y) and non-neural (HeLa) cells and in fibroblasts (from skin biopsies) of patients with homozygous SYNJ1 missense mutation1,2.

Results: We observed that the loss of Synj1 causes a drastic alteration of the early endosomes labeled with EEA1 antibody, which result expanded and more numerous; while it does not affect the morphology of late endosomes labelled with Rab7 antibody both in HeLa and SH-SY5Y cells. In contrast, the morphology of exocytic organells is comparable to wild-type cells. We observed a slight effect on lysosomal compartments stained with a fluorescent dye or with Lamp1 antibody.

Through endocytosis assays we analyzed the trafficking of two receptors (transferrin and EGF), which, once internalized, follow different routes: TfR recycles back to the cell surface, while EGFR move to late endosomes and undergo lysosomal degradation.

Strikingly, in Synj1-depleted cells the TFR trafficking is inhibited with progressive accumulation of transferrin inside the cells. In contrast, we observed no difference in the kinetic of internalization of EGFR in scrambled and silenced cells. Moreover, we found the same alterations in patient fibroblasts: early endosomal compartments result enlarged and the recycling of transferrin is impaired.

Conclusions: Our data indicate that the synaptojanin 1 plays a crucial role in regulating the homeostasis and functions of early endosomal compartments in neural and non-neural cells alterating in particular the recycling pathways from early endosomes. Therefore, we suggest that endosomal pathways dysfunction might underlie neurodegeneration in PARK20.

References: 1: Olgiati S, De Rosa A, Quadri M, Criscuolo C, Breedveld GJ, Picillo M, Pappatà S, Quarantelli M, Barone P, De Michele G, Bonifati V. PARK20 caused by SYNJ1 homozygous Arg258Gln mutation in a new Italian family. Neurogenetics. 2014;15:183-188.

2: Quadri M, Fang M, Picillo M, Olgiati S, Breedveld GJ, Graafland J, Wu B, Xu F,Erro R, Amboni M, Pappatà S, Quarantelli M, Annesi G, Quattrone A, Chien HF,Barbosa ER; International Parkinsonism Genetics Network., Oostra BA, Barone P,Wang J, Bonifati V. Mutation in the SYNJ1 gene associated with autosomal recessive, early-onset Parkinsonism. Hum Mutat. 2013;34:1208-1215.

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MDS Abstracts - https://www.mdsabstracts.org/abstract/alteration-of-early-endosomal-trafficking-causes-neurodegeneration-in-park20/