C9orf72 associates with inactive Rag GTPases and regulates mTORC1‐mediated autophagosomal and lysosomal biogenesis

Abstract

GGGGCC repeat expansion inC9orf72 is the most common genetic cause in both frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS), two neurodegenerative disorders in association with aging. Bidirectional repeat expansions in the noncoding region of C9orf72 have been shown to produce dipeptide repeat (DPR) proteins through repeat-associated non-ATG (RAN) translation and to reduce the expression level of the C9orf72 gene product, C9orf72 protein. Mechanisms underlying C9orf72-linked neurodegeneration include expanded RNA repeat gain of function, DPR toxicity, and C9orf72 protein loss of function. In the current study,we focus on the cellular function of C9orf72 protein. We report that C9orf72 can regulate lysosomal biogenesis and autophagy at the transcriptional level. We show that loss of C9orf72 leads to striking accumulation of lysosomes, autophagosomes, and autolysosomes in cells, which is associated with suppressed mTORC1 activity and enhanced nuclear translocation of MiT/TFE family members MITF, TFE3, and TFEB, three master regulators of lysosomal biogenesis and autophagy. We demonstrate that the DENN domain of C9orf72 specifically binds to inactive Rag GTPases, but not active Rag GTPases, thereby affecting the function of Rag/raptor/mTOR complex and mTORC1 activity. Furthermore, active Rag GTPases, but not inactive Rag GTPases or raptor rescued the impaired activity and lysosomal localization of mTORC1 in C9orf72-deficient cells. Taken together, the present study highlights a key role of C9orf72 in lysosomal and autophagosomal regulation, and demonstrates that Rag GTPases and mTORC1 are involved in C9orf72-mediated autophagy.

C9orf72/ALFA-1 controls TFEB/HLH-30-dependent metabolism through dynamic regulation of Rag GTPases

“…Consistently, in various cellular and animal models, loss of C9orf72 does not cause an overt phenotype under normal conditions but leads to more apparent deficits under stress conditions [18,31,55,56]. Notably, while this manuscript was in review, an independent study reported that C9orf72 associates with Rag GTPases and regulates autophagosomal and lysosomal biogenesis [57], consistent with the main observations of the present study.…”

Cellular and molecular phenotypes ofC9orf72ALS/FTD patient derived iPSC-microglia mono-cultures

“…Likewise, when the number of early endosomes, late endosomes and lysosomes where counted in C9orf72 ALS/FTD iMNs and compared to control iMNs, the loss of lysosomes was the major difference observed between the groups supporting the idea of a potential disruption in the endosomallysosomal pathways and consequently, a contribution to neurodegeneration(107,108). These findings are reinforced by studies reporting enlarged lysosomes in C9orf72 postmortem patient tissues and previous studies proposing that the function of the C9orf72 protein is linked to the regulation of lysosomes and autophagy(30,(130)(131)(132)(133). Our data suggests that similar dysfunctions are present in C9orf72 microglia as shown in our in vitro culture model, but also inC9orf72 ALS/FTD patient postmortem brain tissues.…”

Dysfunctional Autophagy and Endolysosomal System in Neurodegenerative Diseases: Relevance and Therapeutic Options

“…C9Orf72, in a complex with SMCR8 and WDR41, participates in autophagy initiation by activating Rab8a and Rab39b and regulating ULK1, and its loss in neurons leads to the accumulation of protein aggregates (Sellier et al, 2016;Webster et al, 2016;Yang et al, 2016). Puzzling results have shown, however, that C9Orf72 depletion reduces mTORC1 activity and increases lysosomal biogenesis at the transcriptional level, thus pointing to a negative role of C9Orf72 in the autophagic pathway (Ugolino et al, 2016;Wang et al, 2020). Lysosomal deficits, autophagosome accumulation, mitochondrial pathology and impaired retrograde transport of late endosomes have also been reported in the hSOD1 G93A transgenic mouse model.…”