TFEB/Mitf links impaired nuclear import to autophagolysosomal dysfunction in C9-ALS

Abstract

Disrupted nucleocytoplasmic transport (NCT) has been implicated in neurodegenerative disease pathogenesis; however, the mechanisms by which disrupted NCT causes neurodegeneration remain unclear. In a Drosophila screen, we identified ref(2)P/p62, a key regulator of autophagy, as a potent suppressor of neurodegeneration caused by the GGGGCC hexanucleotide repeat expansion (G4C2 HRE) in C9orf72 that causes amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). We found that p62 is increased and forms ubiquitinated aggregates due to decreased autophagic cargo degradation. Immunofluorescence and electron microscopy of Drosophila tissues demonstrate an accumulation of lysosome-like organelles that precedes neurodegeneration. These phenotypes are partially caused by cytoplasmic mislocalization of Mitf/TFEB, a key transcriptional regulator of autophagolysosomal function. Additionally, TFEB is mislocalized and downregulated in human cells expressing GGGGCC repeats and in C9-ALS patient motor cortex. Our data suggest that the C9orf72-HRE impairs Mitf/TFEB nuclear import, thereby disrupting autophagy and exacerbating proteostasis defects in C9-ALS/FTD.

C-Jun N-terminal Kinase Promotes Stress Granule Assembly and Neurodegeneration in C9orf72-mediated ALS and FTD

“…1a ). Furthermore, we show that (G 4 C 2 ) 30 expression in motor neurons using vGlut-GAL4 causes paralysis in pharate flies, as indicated by their inability to eclose (Cunningham et al, 2020). Here, we show that bsk RNAi suppresses this phenotype ( Fig.…”

The Interplay Between Autophagy and RNA Homeostasis: Implications for Amyotrophic Lateral Sclerosis and Frontotemporal Dementia

“…In C9orf72 -FTD/ALS, gain-of-function mechanisms emanating from the repeat are proposed to be the dominant cause of neurodegeneration ( Mizielinska et al, 2014 ; Moens et al, 2019 ), and both repeat RNA and DPRs can affect the autophagy-RNA homeostasis interplay. Dysfunctional nucleocytoplasmic transport from both the repeat RNA and DPRs may impact the nuclear translocation of transcription factors that regulate autophagy genes ( Kim and Taylor, 2017 ; Solomon et al, 2021 ); indeed, the master regulator TFEB is affected by expression of the C9orf72 repeat in Drosophila and cell models, an effect that appeared to be predominantly driven by repeat RNA with only a small effect from poly(GA) ( Cunningham et al, 2020 ). The arginine rich DPRs bind to and disrupt a wide range of membraneless organelles ( Boeynaems et al, 2017 ; Lee K.-H et al, 2016 ), including the nuclear pore through which all nucleocytoplasmic transport occurs, thus may contribute to the aforementioned disruption.…”

“…Furthermore, C9orf72 repeat RNA binds and sequesters several RBPs that have roles in gene regulation (reviewed by McEachin et al, 2020 ). Of note to autophagy, a recent study linked the known C9orf72 repeat RNA and DPR-mediated disruption of nucleocytoplasmic transport ( Kim and Taylor, 2017 ; Solomon et al, 2021 ) to impaired autophagy regulation by TFEB; expression of C9orf72 repeats reduced TFEB translocation to the nucleus resulting in defective autophagy and accumulation of protein aggregates in Drosophila , an effect that appeared to be predominantly driven by repeat RNA, with only a small effect from the DPR poly(GA) ( Cunningham et al, 2020 ) . Consistently, this study described nuclear TFEB depletion in the motor cortex of ALS patients, corroborating a previous study ( Wang et al, 2016 ).…”

Defects of Nutrient Signaling and Autophagy in Neurodegeneration

“…( Shao et al, 2020 ). In Drosophila studies, it has been shown that the repetition of the GGGGCC hexanucleotide impairs the nuclear import of Mitf/TFEB, a transcriptional regulator of autophagolysosomal function, leading to autophagy disruption, accumulation of lysosome-like organelles and proteostasis prior to neurodegeneration ( Cunningham et al, 2020 ).…”