Wednesday, April 2, 2025
4/2/2025
Defining the Mechanisms and Consequences of Nuclear Defects in ALS/FTD - PROJECT SUMMARY A detailed insight into the molecular defects leading to neuronal death in ALS/FTD is still missing, hindering the development of a cure. Defects in the nuclear pore complex (NPC) and nucleocytoplasmic transport (NCT) have been proposed to play a central role in ALS/FTD pathology. An outstanding knowledge gap is what are the mechanisms, consequences, and pathogenic relevance of impaired NCT on neuronal resilience in neurodegenerative diseases and during aging. The objective of this application is to identify at the mechanistic level the novel cellular and molecular pathways that are dysfunctional and cause the failure of the NCT ultimately leading to neurodegeneration. In particular, we will test the hypothesis that the actin cytoskeleton is a main modulator of nuclear stability and NPC function, and that changes to this pathway in ALS/FTD lead to a diminished ability of neurons to cope with stress and physiological stimulation. The specific aims of this proposal are to: 1) Define how actin regulates the function and dysfunction of the NPC. We will perform a systematic and in-depth investigation of the molecular interactions between actin and the NPC using pharmacological and genetic approaches in iPSC-derived neurons carrying the C9ORF72 mutation. 2) Identify the consequences of NCT defects on the nuclear and cytoplasmic proteome in response to cellular stimulation. By combining an “omics” approach with a candidate approach (i.e. CREB pathway), we will analyze changes in the nuclear and cytoplasmic proteome in iPSC-derived neuronal models of ALS/FTD following stimulations, and we will test the potentials of cytoskeleton modulation to rescue such defects. 3) Define how defects in nuclear import affect the transcriptional response to cellular stimulation. We will analyze changes in gene expression and splicing profile in ALS/FTD neurons following cell stimulation by RNA-Seq. Network analysis performed by integrating the results from the transcriptomic and proteomics approaches will identify ALS/FTD relevant altered pathways. Modulation of the cytoskeleton will be used to rescue the identified defects. At the completion of this research, we will have identified the molecular and cellular mechanisms that control the stability and function of the NPC, and the functional consequences that the disruption of NCT has on the resilience of neurons in ALS/FTD. Gaining insights on what pathways upstream and downstream of the NPC are affected in ALS/FTD will greatly expand our understanding of disease pathogenesis and will allow us to identify yet unexplored avenues for therapy in these diseases. Ultimately, we expect that our research will lead to more effective therapeutic strategies that take advantage of the crosstalk between NPC, RNA regulation, and cytoskeleton. This research will have a broad impact on a spectrum of diseases including but not limited to ALS/FTD.
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