Date of Award

January 2014

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Biomedical Sciences

First Advisor

Holly M. Brown-Borg

Abstract

Methylation reactions are important for the establishment and maintenance of epigenetic methylation tags on DNA and histone molecules that are critical for the development and life-long function of an organism. While epigenetics has been studied extensively during embryological development and age-related disease such as cancer, little is known about whether epigenetic methylation enzyme expression change in normally aging or long-living mammals. Because growth hormone (GH) plays an important role in determining the lifespan of many organisms, we have focused our studies on the long-living growth hormone deficient model the Ames dwarf mouse. We focused our studies on epigenetic methylation in the liver, because of its role in GH signaling, IGF-1 signaling, and it's responsiveness to changes in the methionine pathway which govern DNA and histone methylation.

Our studies showed that DNA methyltransferase 1 (DNMT1) protein is basally expressed less in the Ames dwarf compared to their wild-type counterparts, despite higher or equivalent transcription, suggesting that some form of post-transcriptional regulation is taking place. Global DNA methylation levels in dwarf mice resisted age-related changes compared to wild type mice. Histone methylation of key regulatory transcriptional markers, and total histone H3 expression were also altered in dwarf mice.

We demonstrated that the protein abundance of a potential regulatory protein of DNMT1 (DEAD-box helicase 20) significantly higher in Ames dwarf mice at 3, 12, and 24 months of age. Metallothionein1 (MT1) mRNA expression was much higher in dwarf mice especially at 3 months of age. Also there were differences in the methylation of putative CpG islands of the MT1. Growth hormone administration influenced DNA methyltransferase and DDX20 protein expression in dwarf mice and areas of methylation in the predicted MT1 CpG island. MicroRNA 140-3p which pairs with DDX20 for post-transcriptional regulation of DNMT1 was also studied.

Studies using growth hormone receptor knockout mice and over-expressing growth hormone transgenic mice DNMT transcription and interspersed repeat methylation were also studied to discover continuities with previous experiments in Ames dwarf mice. Taken together, this study offers valuable insight about the changes in the epigenetic methylation mechanisms and markers due to aging.

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