Friday, May 9, 2025
5/9/2025
Gene regulatory networks for hair cell regeneration - Project Summary Hair cells are mechanosensory receptors used in the lateral line systems of aquatic vertebrates and in the auditory and vestibular organs of all vertebrates. Only hair cells in zebrafish and in other non-mammalian vertebrates can regenerate completely after being destroyed or damaged by acoustic or chemical exposure. While in mammals, destroying or damaging hair cells results in permanent impairments to hearing and/or balance. To understand how to trigger a regenerative response in humans to repair hearing loss due to the loss of hair cells in the inner ear, it is essential to understand how genes respond to injury and how those responses are controlled in the genome. My research program is focused on building a complete understanding of hair cell regeneration by integrating genomics, high throughput genetics, and phenotypic analysis to investigate gene regulatory networks for hair cell regeneration using the zebrafish as a model system. We previously performed a targeted ablation of the hair cells in adult zebrafish auditory and vestibular organs and interrogated the epigenomic and transcriptomic landscape of regenerating adult inner ear sensory epithelia. Using single-cell analyses, we showed that the support cell population transitions to an intermediate, “progenitor” cell state that become new hair cells and demonstrate that the cell fate decisions may be driven by the coordinate regulation and spatial co-binding of Sox and Six transcription factors. By functionally validating a predicted regeneration responsive enhancer upstream of sox2, we show that precise timing of sox2 expression is critical for hair cell regeneration in zebrafish. We aim to use these findings to gain insight into the molecular mechanisms that regulate hair cell regeneration in zebrafish. To do this, we propose to elucidate how Sox and Six transcription factors control hair cell regeneration. We will examine their spatial and temporal patterns of expression in the regenerating adult inner ear and conduct loss of function studies to assess their roles in hair cell regeneration. Based on chromatin accessibility data on regenerating inner ear tissues at single cell resolution, we have predictions where Sox and Six transcription factors bind. We propose to identify and characterize their direct targets during regeneration using transgenic reporters, CUT&RUN, and CRISPR/Cas9 disruption. We also have a comprehensive dataset of all the enhancer loci activated during hair cell regeneration at cellular resolution. A major component of our research program is identifying new regulators of hair cell regeneration using a mass mutagenesis approach to identify essential enhancers with roles in hair cell regeneration in vivo. These studies will identify a mechanistic understanding of Sox and Six transcription factors in regenerating auditory and vestibular epithelia. We predict that these studies will illustrate the gene regulatory networks involved in hair cell regeneration, which has major implications in discovering therapies to trigger a regenerative response in humans.
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