Mechanisms of disruption of axon transport of autophagic vesicles and lysosomes in C9orf72 ALS

PROJECT SUMMARY
Amyotrophic Lateral Sclerosis (ALS) is a devastating neurodegenerative disease characterized by dying-back
degeneration of upper and lower motor neurons. The most common known cause of familial and sporadic
forms of ALS as well as frontotemporal dementia (FTD) is the GGGGCC hexanucleotide repeat expansion
(HRE) in C9ORF72 (C9). Our laboratory and others recently discovered that impaired nucleocytoplasmic
transport (NCT) is a fundamental and early pathogenic event in C9-ALS that requires stress granule formation.
However, downstream effects of impaired NCT are unclear. Recent studies have shown that autophagosome
biosynthesis occurs in the distal axon followed by retrograde transport of autophagic vesicles (AVs) to the
soma as they mature, providing a potential link between axon transport (AT) and autophagy, two mechanisms
well known to be involved early in ALS pathophysiology. In Drosophila expressing 30 GGGGCC repeats
(30R), we have found an accumulation of p62 and lysosomes, suggesting that impaired regulation of
autophagy and lysosomes may be a pathogenic mechanism for C9-ALS. Further, we have found preliminary
evidence of a reduction in retrograde autophagosome transport in 30R Drosophila. Consistent with this,
preliminary experiments in iPS motor neurons (iPSNs) derived from patients with C9-ALS showed an
accumulation of lysosomes in axons. Specific Aim 1 will further characterize axon transport of multiple cargo in
30R Drosophila and C9 iPSNs using live cell imaging methods. Specific Aim 2 will examine the interrelation
between axon transport, autophagy and lysosomal function and determine if rescuing autophagy can rescue
axon transport deficits of AVs. Finally, preliminary fly data shows that Mitf/TFEB, a transcription factor
regulating autophagy and lysosomes, is mislocalized to the cytoplasm in 30R Drosophila, indicating that
impaired nucleocytoplasmic transport may lead to impaired autophagy and lysosome regulation. Specific Aim
3 will address the hypothesis that impaired nucleocytoplasmic transport is upstream of impaired axon transport
defects and disruptions in autophagy. By using powerful parallel approaches in Drosophila, allowing precise
genetic manipulation of AT and autophagy, and iPSNs derived from patients with C9-ALS, allowing
experimental manipulation of human cells with the disease, this proposal will investigate detailed mechanistic
pathways of axon transport and regulation of autophagy and lysosomes in C9-ALS. Results from these studies
will not only aid our understanding of the pathogenesis and treatment strategies of ALS, but they will also
further our understanding of the axonal biology of autophagy, important in all neurodegenerative diseases.

Grant Number: 5K08NS118123-06
NIH Institute/Center: NIH
Principal Investigator: Sarah Berth