Elucidating competence and fate choice dynamics during retinal development

PROJECT SUMMARY
Degenerative diseases that cause retinal neuronal cell death often result in permanent vision loss. This is
because retinal neurons, like rod and cone photoreceptors, do not regenerate. The most promising potential
therapeutic strategies for restoring lost vision include artificially stimulating endogenous neuronal regeneration
or programming human stem cells into transplantable retinal tissue. However, realizing such strategies is
hindered by our limited understanding of the developmental mechanisms used to build the retina.
During retinal development, proliferative multipotent progenitor cells choose between seven major fate
outcomes. This choice process, called fate specification, is determined by a combination of a cell’s potential (i.e.,
competence) and instructive factors that select between competing fate choices. Retinal fate specification is a
dynamic, probabilistic process that is controlled by the intersection of intrinsic gene regulatory networks and
environmental cell-cell signaling mechanisms. The Notch signaling pathway impacts competence and fate choice
decisions in the retina. However, its mechanisms are poorly understood due to a lack of genetic tools that can
dynamically manipulate signaling in specific subpopulations of competent cells over time. To overcome this
barrier, we identified enhancer sequences for the key transcription factor Otx2 that drive discrete spatial and
temporal activity patterns in the mouse retina. Using these narrowly tailored enhancer tools, our initial findings
show that Notch signaling plays multiple discrete fate choice roles throughout development. Our objective is to
finely dissect how Notch signaling functions to understand the probabilistic nature of retinal cell fate specification.
In Aim 1 of this proposal, we will investigate how Notch signaling regulates multiple different fate decisions
throughout retinal development. We will activate Notch signaling at discrete stages of retinal development and
use single cell RNA sequencing and histological approaches to determine how cells change competence and
fate choices over time. These data will be used to determine whether Notch signaling delays decision making or
acts by specifically instructing fate choices in competent cells at different stages of development. In Aim 2, we
will use developmental and genetic techniques to explore how Notch signaling exposure (dosage and duration)
differentially impacts competence and retinal cell fate decisions.
Leveraging our unique genetic tools, this project will reveal how the multifaceted Notch signaling pathway
impacts competence and dynamic fate choice probabilities in the developing retina. This knowledge is essential
for creating regenerative and cell-based therapies to replace lost neurons, which may restore vision in millions
of people suffering from retinal degenerative diseases.

Grant Number: 1R21EY037026-01A1
NIH Institute/Center: NIH
Principal Investigator: Joseph Brzezinski