Date of Degree

6-2021

Document Type

Dissertation

Degree Name

Ph.D.

Program

Biology

Advisor

Mark Emerson

Committee Members

Bao Vuong

Daniel Weinstein

Carol Mason

Ruth Johnson

Subject Categories

Biology | Cell Biology | Developmental Biology | Molecular Genetics

Abstract

During development, retinal progenitor cells (RPCs) divide to form all of the cell types that make up the retina. Multipotent RPCs are competent to generate all retinal cell types, while restricted RPCs form specific lineages of cells. In particular, one genetically-defined RPC type preferentially gives rise to cone photoreceptors and horizontal cells. Many of the mechanisms that are responsible for directing cell fate choice within this lineage are unknown. This thesis largely focuses on examining the development of specific cell types and subtypes from restricted RPCs and on investigating the gene regulatory events that underlie cone photoreceptor and horizontal cell development in the retina.

Short-term assays have concluded that ThrbCRM1 RPCs preferentially generate cone photoreceptors and horizontal cells. However, developmental timing has precluded an extensive cell type characterization of their progeny. The first part of this thesis describes the development and validation of a recombinase-based lineage tracing system for the chicken embryo to further characterize the lineage of these cells. ThrbCRM1 RPCs were found to preferentially form photoreceptors and horizontal cells, as well as a small number of retinal ganglion cells. The photoreceptor cell progeny are exclusively cone photoreceptors and not rod photoreceptors, confirming that ThrbCRM1 RPCs are restricted from the rod fate. In addition, specific subtypes of horizontal cells and retinal ganglion cells were overrepresented, suggesting that ThrbCRM1 RPCs are not only restricted for cell type, but for cell subtype as well.

The second part of this thesis utilizes the identification of novel cis-regulatory elements as a method to examine the gene regulatory networks that direct the development of horizontal cells. The OC1ECR22 and Tfap2aACR5 elements were shown to be enhancers for Onecut1 (OC1) and Tfap2a, respectively, and to be specifically active in developing horizontal cells. The OC1ECR22 element is activated by Ptf1a and Rbpj, which translates to regulation of OC1 expression and suggests that Ptf1a is a direct activator of OC1 expression in developing horizontal cells. The region within the Tfap2aACR5 element that is responsible for its activation was determined to be a 100 bp sequence named Motif 4. Both OC1ECR22 and Tfap2aACR5 are negatively regulated by the nuclear receptors Thrb and Rxrg, as is the expression of OC1 and Tfap2a, suggesting that nuclear receptors may have a role in the negative regulation of horizontal cell development.

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