Methods and Mechanisms of DNA Methylation in Development and Disease

<p>DNA methylation is a mechanism for long-term transcriptional regulation and is required for normal cellular differentiation. Failure to properly establish or maintain DNA methylation patterns leads to cell dysfunction and diseases such as cancer and neurological disorders. The goal of this thesis is to understand the role of DNA methylation in oncological cellular transformation and in normal development. To achieve this goal, I have developed a novel method for mapping genome-wide DNA methylation patterns and have applied the method to gonadectomy-induced adrenocortical neoplasms and to maturing motor neurons. The novel method, called Laser Capture Microdissected-Reduced Representation Bisulfite Sequencing (LCM-RRBS), accurately and reproducibly profiles genome-wide methylation of DNA extracted from microdissected fresh frozen or formalin-fixed paraffin-embedded tissue samples. Using this method, I find that significant DNA methylation changes, associated with attendant expression changes, occur in transformed adrenocortical cells. My work has also uncovered significant DNA methylation configuration in maturing motor neurons associated with dramatic expression changes. I show that demethylated regions are enriched for known neuron-specific transcription factor binding sites and that genetic disruption of the active demethylation machinery significantly inhibits motor neuron differentiation and maturation. Together, these experiments demonstrate that DNA methylation plays a role in the transformation of normal cells to cancer cells and that DNA methylation is critical to proper motor neuron formation. I conclude that aberrant DNA methylation controls gene expression in gonadectomy-induced adrenocortical neoplasms and that neuron-specific transcription factors could recruit demethylating enzymes to regions that lose DNA methylation in motor neurons upon maturation.</p>

Name thesis:degree_name

Doctor of Philosophy (PhD)

Level thesis:degree_level

Dissertation

Discipline thesis:degree_discipline

Biology and Biomedical Sciences: Molecular Genetics and Genomics

Year dc:date.available

2014

Author dc:creator

• Schillebeeckx, Maximiliaan

Contributors dc:contributor

• Robi D Mitra

• Adrenal
• Bisulfite Sequencing
• DNA Methylation
• Motor Neurons

Language dc:language

English (en)

DOI

https://doi.org/10.7936/K73B5X5X

OAI identifier oai:identifier

oai:openscholarship.wustl.edu:etd-2343