Horstick, E. | Search bioRxiv

bioRxiv2023

Molecular regulation of retinal regeneration is context specific

Many genes are known to regulate retinal regeneration following widespread tissue damage. Conversely, genes controlling regeneration following limited retinal cell loss, akin to disease conditions, are undefined. Combining a novel retinal ganglion cell (RGC) ablation-based glaucoma model, single cell omics, and rapid CRISPR/Cas9-based knockout methods to screen 100 genes, we identified 18 effectors of RGC regeneration kinetics. Surprisingly, 32 of 33 previously known/implicated regulators of retinal tissue regeneration were not required for RGC replacement; 7 knockouts accelerated regeneration, including sox2, olig2, and ascl1a. Mechanistic analyses revealed loss of ascl1a increased "fate bias", the propensity of progenitors to produce RGCs. These data demonstrate plasticity and context-specificity in how genes function to control regeneration, insights that could help to advance disease-tailored therapeutics for replacing lost retinal cells. One sentence summaryWe discovered eighteen genes that regulate the regeneration of retinal ganglion cells in zebrafish.

neuroscience↗

bioRxiv2022

The Musashi proteins are post-transcriptional activators of protein expression and alternative exon splicing in vertebrate photoreceptors

The Musashi proteins, MSI1 and MSI2, are conserved RNA binding proteins with a role in the maintenance and renewal of stem cells. Contrasting with this role, retina and terminally differentiated photoreceptor cells express high levels of MSI1 and MSI2, indicating that the two proteins have a role unrelated to maintaining undifferentiated cell state. Here we show that the Musashi proteins are essential in mature photoreceptors. Combined knockout of Msi1 and Msi2 lead to loss of the retina response to light and progressive photoreceptor cell death. The two proteins are fully redundant as individual deletion of Msi1 or Msi2 did not produce a phenotype. To define the molecular functions underlying the requirement for Musashi in photoreceptors, we delineated their RNA targets and analyzed the effect of the combined Msi1/Msi2 knockout on transcript levels, pre-mRNA splicing, and protein expressions. We show distinct nuclear and cytoplasmic functions for the Musashi proteins in photoreceptor cells. In the nucleus, Musashi binding to the downstream proximal intron promotes splicing of alternative exons. Surprisingly, four conserved photoreceptor-specific alternative exons in genes critical for vision proved to be dispensable, leaving open questions about the selective pressures that lead to the conservation of these exons and the contribution of alternative splicing to the phenotype of the Musashi knockout. In the photoreceptor cell cytoplasm MSI1 and MSI2 act as activators of protein expression. The combined knockout of Msi1 and Msi2 reduced the levels of multiple proteins including proteins required for vision and photoreceptor survival.

neuroscience↗