Mechanisms of impaired ESCRT-III nuclear surveillance in ALS/FTD

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Amyotrophic Lateral Sclerosis (ALS) and Frontotemporal Dementia (FTD) are two age-related neurodegenerative diseases that share genetic and pathological underpinnings. Recently, disruptions to the composition and function nuclear pore complex (NPC) have been identified as a prominent and early pathomechanism underlying neurodegenerative diseases including ALS, AD/FTD, and HD. As the NPC and surrounding nuclear envelope (NE) environment critically control fundamental cellular processes including nucleocytoplasmic transport (NCT), gene expression, and genome organization, NPC and NE homeostasis is essential for neuronal function and survival. Using induced pluripotent stem cell (iPSC) derived neurons (iPSNs) and postmortem human CNS tissues, we have recently identified a significant injury to the NPC characterized by the reduction in specific Nups from the human neuronal NPC that culminates in impaired nuclear protein import and contributes to the dysfunction and mislocalization of the RNA binding protein TDP- 43, a prominent pathological event in neurodegeneration. This NPC injury cascade is initiated by the aberrant nuclear accumulation of CHMP7 in sALS and C9orf72 ALS/FTD neurons. CHMP7 is an ESCRT-III protein that while normally localized predominantly to the cytoplasm, is thought to function primarily in ESCRT-III mediated nuclear surveillance events. The cell biological events and molecular mechanisms underlying CHMP7’s contribution to NPC and NE surveillance and homeostasis are beginning to be understood in yeast and non- neuronal mammalian cells during cell division, NE breakdown, and NPC insertion. However, little is understood regarding CHMP7’s role in maintaining NPC and NE homeostasis in long-lived, non-dividing human neurons where NPC components are infrequently exchanged, and NE breakdown does not routinely occur. Our preliminary data highlight the fact that the molecular events that regulate the nuclear localization and activation of CHMP7 mediated nuclear surveillance in neurons are not perfectly concordant with those recently identified in yeast and dividing mammalian cells suggestive of distinct neuronal mechanisms that give rise to CHMP7 initiated pathogenic cascades in ALS/FTD. In the proposed studies, we will employ iPSNs from a large number of sALS and C9orf72 ALS/FTD patients and appropriate controls and utilize a combination of candidate based siRNA knockdown, biochemical protein interaction techniques, recently developed and optimized live cell and super resolution imaging strategies, and expression of a newly generated CHMP7 variant plasmid library. Collectively, these studies will identify the molecular mechanisms that facilitate the nuclear entry (Aim 1) and nuclear export (Aim 2) of CHMP7 as well as the initiation of CHMP7 mediated NPC injury (Aim 3) to give rise to NPC injury cascades in ALS/FTD. Therefore, these studies will provide an essential understanding of the cell biological and molecular events that facilitate CHMP7 mediated NPC injury in human neurons and provide insights into therapeutic strategies for alleviating this early and significant pathogenic cascade in ALS/FTD.