Patterning Genes in Retinal Development – Project Summary
The long-range goal of this grant is to determine the cellular and molecular events that lead to the
generation of specific cell types in the vertebrate retina. In this renewal we focus upon mechanisms regulating
differential expression of the cone visual pigment genes on tandemly-replicated gene arrays. In humans
a tandemly-replicated array on the X chromosome consists of one long wavelength-sensitive (LWS) gene
followed by 1-9 medium wavelength-sensitive (MWS) genes, which have diverged spectrally from LWS. This
recent replication has provided most humans with trichromatic color vision, because LWS, MWS, and SWS1
opsins are uniquely expressed in separate cone populations. Heritable defects in the LWS/MWS array result in
various forms of color blindness, X-linked retinal degenerations, and Bornholm Eye Disease, a cone
dysfunction associated with high myopia. Insights into regulation of differential expression of LWS vs. MWS
opsins could allow therapeutic manipulation of gene expression to treat these disorders. In addition, future
regenerative approaches to the treatment of other retinal disorders that involve the loss of cones (age-related
macular degeneration; Stargardt’s disease) would ideally include similar in vitro or in vivo manipulations to
generate cone phenotypes in ratios that support high-acuity color vision.
The widely-accepted model for human LWS vs. MWS opsin regulation states that a stochastic event
favors an association of an upstream regulatory region with the LWS or most proximal MWS. However,
topographic patterns of the LWS:MWS ratio suggest that a nonrandom, trans regulatory mechanism may be
involved. Pursuit of regulatory mechanisms has been challenging because within mammals, only primate
genomes contain tandem opsin arrays. In contrast, the genomes of teleost fish, including zebrafish, contain
numerous tandem arrays of opsins, which are the consequences of independent gene replication events and
neofunctionalization. In our published and preliminary data we demonstrate that in zebrafish, the
developmental signaling molecules retinoic acid (RA), and thyroid hormone (T3) can each control differential
expression of the tandem duplicates, LWS1 vs. LWS2, an array orthologous to the human LWS/MWS array.
Furthermore, compelling preliminary data suggest that RA can promote expression of LWS opsin in human
iPSC-derived 3D retinal organoids, changing the ratio of LWS:MWS. Together these findings lay the
groundwork for a tremendous breakthrough in understanding determination of LWS vs. MWS cone subtype.
In this renewal we pursue mechanisms through which RA and T3 control differential expression of
tandemly replicated opsins, and apply this knowledge to retinal regeneration and human retinal organoid
development, with three Specific Aims: 1. Determine the relative roles of RA and T3 signaling and their
receptors as endogenous regulators of differential expression of tandemly replicated opsin genes. 2.
Determine mechanisms through which RA and T3 signaling regulate tandemly replicated opsin genes. 3.
Determine roles of RA and T3 signaling for controlling cone fates during retinal regeneration and in human
iPSC-derived 3D retinal organoids. These studies will uncover novel mechanisms for the differential expression
of tandemly replicated opsin genes, and will generate information necessary to manipulate cone phenotypic
fates in concert with the application of regenerative or cell replacement therapies for human retinal