# CRX AND ITS REGULATORY NETWORK IN RETINAL DEGENERATIONS

> **NIH NIH R01** · WASHINGTON UNIVERSITY · 2020 · $518,466

## Abstract

PROJECT SUMMARY
 Photoreceptor development and maintenance require precisely regulated gene expression. This
regulation is controlled by the Gene Regulatory Network (GRN) of photoreceptor transcription factors (TFs)
and their target non-coding cis-regulatory DNA elements (CREs). Mutations in either TFs or CREs can cause
misregulation, leading to photoreceptor diseases. How mutations in coding sequences cause diseases, such
as CRX-linked retinopathies, has been extensively studied, and many such mutations have been identified and
can be tested for. However, only a few disease-associated mutations in non-coding DNA regulatory elements
are known. One example is X-linked blue-cone monochromacy, a red/green color blindness associated with
disruption of the Locus Control Region (LCR), a distal CRE of the Red/Green gene array. Little is known about
the location or functional importance of other retinal CREs, presenting a bottleneck for identifying and
evaluating disease-associated variants in the largely unexplored non-coding portion (99%) of the genome.
Thus, there is an urgent need to catalog functional CREs and understand their sequence logic. To this end, we
have identified the subset of CRX bound CREs that are Dependent on the activity of CRX to establish an
active chromatin state and promote photoreceptor gene expression. We hypothesize that many of these
CREs are essential for rod gene expression, cell fate, and survival, and their sequence features
determine how they mediate the commands of CRX and/or other TFs. We propose to test this hypothesis
in three specific aims using innovative high-throughput functional genomics approaches. Aim 1 will provide a
deep understanding of two previously-identified CREs using loss-of-function studies in mice: By thoroughly
characterizing the individual and combined deletions of the two rhodopsin (Rho) enhancers CBR and RER, we
will decipher their in vivo roles in regulating endogenous Rho expression, rod cellular function, and health.
Aims 2 and 3 will identify the subset of essential retinal CREs using unbiased high-throughput functional tests
in mouse retinas: We will first identify activating CREs that are sufficient to enhance the expression of a
reporter gene driven by a minimal rod gene promoter using massively parallel reporter assays (MPRA) (Aim
2); We will then identify active CREs that are necessary for rod gene expression and cell identity using a
CRISPR-Cas9 genomic deletion screen (Aim 3). Finally, we will combine the information gained from these
two complementary approaches to decipher the sequence logic mediating CRE regulatory function. Ultimately,
we will identify the most functionally important photoreceptor CREs, gain a deep understanding of their genetic
and epigenetic grammar, and be able to predict the effects of specific disruptions to these CREs on rod gene
expression and cell fate. The findings will significantly advance our understanding of normal and pathogenic
photoreceptor gene ex...

## Key facts

- **NIH application ID:** 9852448
- **Project number:** 5R01EY012543-20
- **Recipient organization:** WASHINGTON UNIVERSITY
- **Principal Investigator:** SHIMING CHEN
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $518,466
- **Award type:** 5
- **Project period:** 2000-03-01 → 2023-01-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9852448

## Citation

> US National Institutes of Health, RePORTER application 9852448, CRX AND ITS REGULATORY NETWORK IN RETINAL DEGENERATIONS (5R01EY012543-20). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/9852448. Licensed CC0.

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