Abstract

Lmx1b, a LIM homeodomain transcription factor is necessary for limb, kidney, eye, and central nervous system development. Lmx1b knockout (KO) mice display; abnormal limb dorsalization with distal ventral-ventral limbs, nephropathy, ocular malformations, loss of dopaminergic/serotonergic neurons, and reduced cerebellum and tectum formation. The developmental mechanisms utilized by Lmx1b remain largely unknown; to investigate downstream targets, we have used the limb as a model because of its accessibility and striking Lmx1b KO phenotype. To identify candidate genes, we compared gene arrays from Lmx1b KO and wildtype mouse limbs during limb dorsalization, i.e., 11.5, 12.5, and 13.5 days post coitum. We identified 54 target genes differentially expressed in all three stages. An abundance of extracellular matrix-related targets were found to be regulated by Lmx1b, including collagens and proteoglycans. Three Lmx1b-regulated proteoglycans are clustered together at one genomic locus, Keratocan, Lumican, and Decorin (KLD). To determine whether Lmx1b's regulation of these targets is direct, we used in silico comparative genomic analysis and identified a potential regulatory region with a known Lmx1b binding site (Peak 3). We hypothesized that Lmx1b directly regulates KLD via Peak 3. We isolated Peak 3 and linked it to a green fluorescent protein (GFP) reporter. This construct was electroporated into chick embryos at the primitive streak stage (Hamburger-Hamilton (HH) 4) and during early neural tube formation (HH10). GFP expression was detected in neural tube and surrounding the developing midbrain and cerebellum. Expression of Lmx1b and KLD mRNA overlaps the activity of this regulatory region. Our data suggests Lmx1b regulates morphogenesis by altering the extracellular matrix composition such as KLD proteoglycans. Furthermore, the activity of KLD-associated Peak 3 localized to Lmx1b-dependant CNS structures during development supports a direct role for Lmx1b regulation.

LLU Discipline

Anatomy

Department

Basic Sciences

School

School of Medicine

First Advisor

Oberg, Kerby C.

Second Advisor

Fallon, John: Payne, Kimberly

Third Advisor

Subburaman, Mohan

Fourth Advisor

Wall, Nathan

Degree Name

Doctor of Philosophy (PhD)

Degree Level

Ph.D.

Year Degree Awarded

January 2012

Date (Title Page)

6-1-2012

Language

English

Library of Congress/MESH Subject Headings

Central Nervous System -- Growth and Development; Embryonic Development; Kidneys -- Growth and Development; Eye -- Growth and Development; Extremities -- Growth and Development

Subject - Local

Limb Dorsalization; Central Nervous System Development; LIM Homeodomain Transcription Factor; Kidney Development; Eye Development

Type

Dissertation

Page Count

190 p.

Digital Format

Application/PDF

Digital Publisher

Loma Linda University Libraries

Usage Rights

This title appears here courtesy of the author, who has granted Loma Linda University a limited, non-exclusive right to make this publication available to the public. The author retains all other copyrights.

Collection

Loma Linda University Electronic Theses & Dissertations

Collection Website

http://scholarsrepository.llu.edu/etd/

Repository

Loma Linda University. Del E. Webb Memorial Library. University Archives

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