A genetic program for organ regeneration in Zebrafish

Stowers Institute For Medical Research, Kansas City MO

Investigators

Abstract

Project Summary Sensory hair cells in the mammalian inner ear and vestibular system are respectively responsible for hearing through the transduction of air vibrations to sound, and the detection of gravity and providing balance. Once they are damaged or killed, sensory hair cells do not regenerate, leading to permanent hearing loss and vestibular disfunction. Despite research efforts, regeneration of mammalian hair cells is limited, and hearing and vestibular function have never been restored. Thus, there is a pressing need to further investigate hair cell regeneration. We have identified zebrafish as an excellent model to address this need. Sensory hair cells of mammals and zebrafish are functionally and genetically homologous, and hair cells in zebrafish rapidly regenerate and exist in a state of constant turnover. Zebrafish are an established research organism, they are genetically tractable with mutation and transgenesis, and rapid regeneration of hair cells is ideal for performing assays. A number of genes are known to influence hair cell regeneration in zebrafish, but how they interact in a gene regulatory network (GRN) is still unknown. I believe that understanding the global epigenetic landscape and complex genetic regulatory interactions between genes is essential to making progress on hair cell regeneration. My objective is to assemble a GRN describing the genetic and regulatory interactions that drive hair cell regeneration, and to test the GRN by manipulating individual components. I hypothesize that the loss of regeneration in mammals is caused by evolutionary changes to the hair cell GRN. This project will define the regulatory interactions driving hair cell regeneration in zebrafish and identify specific genes and components of the GRN that are essential for this process as targets for mammalian studies. I will characterize the global regulatory landscape using ChIP-seq and ATAC-seq in a fine time scale during hair cell regeneration and in homeostasis. I will then perform a bioinformatic analysis to build a GRN integrating ATAC-seq and ChIP-seq with scRNA-seq data. I will search for enriched transcription factor binding motifs associated with genes changing during hair cell regeneration, and use these enriched motifs to draw genetic connections between transcription factors and target genes. Lastly, I will test GRN connections by mutating genes and enhancers involved in regeneration, and assay phenotypes to determine the key genes driving the process. I will clone enhancers of orthologous genes from mice into zebrafish to determine where the GRN has changed between the two species, and attempt to restore connections by adding transcription factor binding back to mouse enhancers. My rigorous PhD training has prepared me to carry out this research project and expand upon my previous experience testing GRNs and carrying out mutagenesis and phenotypic assays. With the support of core facilities, I have already generated high quality preliminary ATAC-seq and ChIP-seq data, used it to find new GRN connections, and demonstrated regeneration phenotypes by identifying and mutating key regeneration genes. Through this research, I expect to make significant contributions towards understanding hair cell regeneration, providing new gene and regulatory targets to be used in mammalian studies and eventually therapeutic interventions.