Arf6 and the 5'phosphatase of Synaptojanin 1 regulate autophagy in cone photoreceptors.

Ashley A George,Susan E Brockerhoff,Sara Hayden,Gail R Stanton

doi:10.1002/icl3.1044

Abstract

Abnormalities in the ability of cells to properly degrade proteins have been identified in many neurodegenerative diseases. Recent work has implicated Synaptojanin 1 (SynJ1) in Alzheimer's disease and Parkinson's disease, although the role of this polyphosphoinositide phosphatase in protein degradation has not been thoroughly described. Here we dissected in vivo the role of SynJ1 in endolysosomal trafficking in zebrafish cone photoreceptors using a SynJ1-deficient zebrafish mutant, nrca14 . We found that loss of SynJ1 leads to specific accumulation of late endosomes and autophagosomes early in photoreceptor development. An analysis of autophagic flux revealed that autophagosomes accumulate due to a defect in maturation. In addition we found an increase in vesicles that are highly enriched for PI(3)P, but negative for an early endosome marker in nrca14 cones. A mutational analysis of SynJ1 enzymatic domains found that activity of the 5' phosphatase, but not the Sac1 domain, is required to rescue both aberrant late endosomes and autophagosomes. Finally, modulating activity of the PI(4,5)P2 regulator, Arf6, rescued the disrupted trafficking pathways in nrca14 cones. Our study describes a specific role for SynJ1 in autophagosomal and endosomal trafficking and provides evidence that PI(4,5)P2 participates in autophagy in a neuronal cell type.

Highlights

Cells must balance synthesis of new proteins with the degradation of old and damaged proteins to maintain their cellular proteome
To correlate the endolysosomal defects observed in nrca14 cones [18] with initial stages of photoreceptor development, we examined late endosomes and autophagosomes in cone photoreceptors starting at 3dpf
These results indicate that Arf6a and Synaptojanin 1 (SynJ1) are acting in the same pathway to modulate autophagy in cone photoreceptors

Summary

Introduction

Cells must balance synthesis of new proteins with the degradation of old and damaged proteins to maintain their cellular proteome. Many neurodegenerative diseases including Alzheimer’s, Parkinson’s, and Amyotrophic Lateral Sclerosis show cellular phenotypes that are consistent with abnormalities in protein turnover[1]. Protein degradation is accomplished by two main pathways; the ubiquitin-proteasome system and the endolysosomal system. Proteins destined for degradation will continue through the endolysosomal pathway, first through late endosomes and to the lysosome. Cytosolic proteins can be degraded either by the proteasome or by autophagy[3]. The term autophagy generally refers to macroautophagy, a process in which a double membrane structure forms in the cytoplasm, non- engulfing cytoplasmic contents including proteins and organelles. In order to degrade their contents, autophagosomes must fuse with the lysosome. Autophagosomes can fuse directly with the lysosome or first fuse with other endosomal compartments, forming an amphisome, before fusing with the lysosome[4]

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