Investigating non-coding RNA 7SK in Huntington's Disease neurodegeneration using direct neuronal conversion

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PROJECT SUMMARY Aging is the greatest risk factor across neurodegenerative diseases and should be recapitulated in adult-onset disease research models. However, current models utilize human neurons differentiated from induced pluripotent stem cells, which erase cellular signatures of aging. The Yoo Laboratory has pioneered a system that uses ectopic expression of microRNAs-9/9* and -124 (miR-9/9*-124) to directly convert human adult fibroblasts (HAFs) into microRNA-induced neurons (miNs). The miNs maintain molecular age-associated properties, including the epigenetic clock, telomere lengths and oxidative stress signatures. Additional transcription factors can synergize with miR-9/9*-124 to generate neuronal cell subtypes including microRNA- induced medium spiny neurons (MSNs), the primary cells lost in Huntington’s Disease (HD). These subtypes can model cellular pathologies in adult-onset neurodegenerative disease, such as endogenous aggregation, DNA damage, mitochondrial dysfunction, and cell death. Degenerating neurons in multiple disorders, including HD, also demonstrate increased long gene expression (LGE), chromatin dysregulation, and hyperexcitability in comparison to healthy neurons, yet these HD-associated features have not been successfully targeted for treatment. This proposal builds on my published first-author study showing that the small nuclear RNA RN7SK (7SK) is required for neuronal chromatin accessibility and transcription activation across the genome during miR-9/9*-124 mediated reprogramming. A substantial portion of these neuronal loci and LGE correspond to genes that are dysregulated in HD-MSNs compared to control MSNs. Here, I propose to use my previous findings to study how depletion of 7SK expression in HD-MSNs can possibly improve HD neurodegenerative phenotypes. In Aim 1, I plan to test if knocking down 7SK can bring LGE in HD-MSNs to control levels and if it can modulate HD-associated chromatin signatures. Following in Aim 2, I propose to assess if repression of 7SK can ameliorate HD-associated neurodegenerative phenotypes, including hyperexcitability, metabolic dysfunction, and spontaneous cell death. Combining molecular, genomic, electrophysiological, and cell pathology assays, I will evaluate if reduction of 7SK can restore these HD-associated molecular features and cellular phenotypes to healthy control MSN levels. Completion of these aims will reveal implications of neuronal chromatin and transcriptional regulation for alleviating HD-associated phenotypes. These data will provide foundational knowledge for the advancement of adult-onset neurodegenerative disease modeling and explore putative epigenomic targets for HD therapeutic development.