Author's School

Graduate School of Arts & Sciences

Author's Department/Program

Biology and Biomedical Sciences: Molecular Cell Biology

Language

English (en)

Date of Award

Spring 3-9-2014

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Chair and Committee

Joseph Corbo

Abstract

The transcription factor neural retina leucine zipper (Nrl) is a critical determinant of rod photoreceptor fate and a key regulator of rod differentiation. Nrl(-/-) rod precursors fail to turn on rod genes and instead differentiate as cones. Furthermore, NRL mutations in humans cause retinitis pigmentosa, a potentially debilitating disease. Despite the developmental and clinical significance of this gene, however, little is known about the transcriptional regulation of Nrl itself. In this study, we sought to define the cis- and trans-acting factors responsible for initiation and maintenance of Nrl transcription in the mouse retina. Utilizing a quantitative mouse retinal explant electroporation assay, we discovered a phylogenetically conserved, 30-base pair region immediately upstream of the transcription start site that is required for Nrl promoter activity. This region contains critical binding sites for the retinal transcription factors Crx, Otx2, and ROR-beta, and experiments demonstrate binding of these factors both in vitro and in vivo. We propose a model in which Nrl expression is primarily initiated by Otx2 and ROR-beta and later maintained at high levels by Crx and ROR-beta.

In addition to elucidating the developmental regulation of Nrl, we sought to determine whether manipulation of this photoreceptor cell fate switch might be applicable to the treatment of heritable blindness. Retinitis pigmentosa is one of the most common degenerative diseases of the eye and is associated with early rod photoreceptor death followed by secondary cone degeneration. We hypothesized that converting adult rods into cones would make the cells resistant to the effects of mutations in rod-specific genes, thereby preventing secondary cone loss. To test this idea, we engineered a tamoxifen-inducible allele of Nrl to acutely inactivate the gene in adult rods. This manipulation resulted in reprogramming of rods into cells with cone-like molecular, histologic and functional properties. Moreover, reprogramming of adult rods achieved cellular and functional rescue of retinal degeneration in a mouse model of retinitis pigmentosa. These findings suggest that elimination of Nrl in adult rods may represent a novel therapy for retinal degeneration.

Comments

Permanent URL: http://dx.doi.org/10.7936/K7WW7FP8

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