Research Experience
InvestiGators, Dr. Peggy R. Borum’s Lab, University of Florida
My research career began in a metabolic biochemistry lab run by Dr. Peggy Borum. Dr. Borum’s research centers on metabolism during in normal systems as well as in patients with HIV or epilepsy. The molecule that binds her work together is the amino acid derivative carnitine. One of carnitine’s roles in the cell is to transport fatty acids across the mitochondrial membranes so that they can be hydrolyzed for energy (beta-oxidation) or utilized for particular purposes. By measuring carnitine and which fatty acids it is bound to, researchers can determine a patient’s metabolic state (Thompson, et al. 2012), potentially leading to more specific treatments.
As an undergraduate, I joined Dr. Borum’s team to gain experience with research with direct clinical impact. The lab was then creating a model of neonatal carnitine metabolism using piglets. These week-long experiments required the coordination of more than a dozen individuals to care for the animals, collect samples, and prepare the samples for analysis. Similarly, we regularly collected blood samples from epileptic patients who were placed on a high-fat, ketogenic diet.
My personal project focused on how metabolic and hormonal perturbations affected carnitine concentrations in the blood and organs of rats. I found that inducing diabetes and altering the sex hormones of these rats greatly affected how carnitine was distributed in the body. Furthermore, carnitine concentrations in individual organs and the blood did not all change in the same manner. It is a long-held assumption that measuring blood carnitine concentrations is indicative of whole-body carnitine levels, but our results indicated that the real situation is much more complicated.
Through work with Dr. Borum, I gained valuable experience with experimentation, biochemistry, statistics, and systems biology. Through administrative work with the InvestiGators, I began to appreciate the organization and regulation required in a research lab. I presented posters at the Experimental Biology conference and co-authored a review paper on the role of carnitine in brain physiology (Jones, et al. 2010). Above all else, Dr. Borum sparked my curiosity in academic research and biology that inspired me to go to graduate school.
Dr. Douglas A. Marchuk’s Lab, Duke University
I was accepted to graduate school at Duke University through the Cell and Molecular Biology Program. After completing my rotations, I joined the lab of Dr. Doug Marchuk due to his reputation for mentorship and my enjoyment of his projects in human genetics. He gave me a project to study a human disease called cerebral cavernous malformations (CCM) in which some capillaries in the brain grow to the point where they become leaky, leading to seizures, numbness, stroke, and death.
Mutations in one of three genes have been found to cause CCM (the genes were thus called CCM1, CCM2, and CCM3). Based on this information, Dr. Marchuk’s lab created the original mouse models of CCM1 and CCM2. In order to produce more CCMs in these mice, a sensitizer mutation in Trp53 (encoding the protein p53, known as the guardian of the genome) was introduced that increase the rate of somatic mutations, thereby increasing the number of CCM lesions in CCM1 and CCM2 mice.
My first project in the lab was to improve these mouse models so that we could begin testing new therapies. One problem with using Trp53 as a sensitizer mutation is that it can cause various types of cancer in the mice that prevent long-term study. I used mutations in Msh2, part of the mismatch repair complex, instead to create newer mouse models that lived for longer than their predecessors (McDonald, et al. 2011). We also noted that brain vessels in these mice displayed high activity of Rho Kinase (ROCK). Next, we treated the mice with an inhibitor of ROCK called fasudil, and found a large decrease in the numbers of CCM lesions (McDonald, et al. 2012). Subsequent experiments will need to determine if fasudil or ROCK inhibition in general is a viable treatment for CCM patients.
While at Duke, I also studied the genetics of how CCMs are formed. Previous work in the lab focused on patients with a family history of CCM (inherited cases). In these cases, a CCM mutation is inherited from the mother or father, and a subsequent, somatic mutation occurs in the brain vasculature to the remaining wild-type copy of the CCM gene (Akers, et al. 2009). No one had looked closely at non-inherited (sporadic) cases, however, so that was my next project.
Using next-generation sequencing, I was able to show that sporadic CCM lesions also contain somatic mutations in one of the three CCM genes, including a case where I found two biallelic, somatic mutations within the same lesion (McDonald, et al. 2014). Such evidence indicates that both inherited and sporadic cases of CCM have the same underlying genetic mechanism. Therefore, both types may be treatable by the novel therapies in development.
Dr. Jodie M. Fleming’s Lab, North Carolina Central University
While completing my dissertation at Duke, I began learning how to use different computational methods to analyze large data sets. In looking for postdoctoral research opportunities, I sought out a project where I could learn and apply further bioinformatics techniques. Dr. Fleming studies molecular mechanisms underlying breast cancer, and she had a large data set to examine gene expression of epithelial cells, adipocytes, and fibroblasts from normal breast tissue. After cleaning up and normalizing the data, I employed principal components analysis to identify which genes in the data set were having the largest effect. Subsequent analysis has focused on finding connections between various clinical variables and the expression of these genes in the different cell types.
One of the genes previously identified in breast cancer studies is lipolysis-stimulated lipoprotein receptor (LSR). I found that LSR has nine predicted transcript variants that may each have separate biological effects. I employed computational techniques to predict the structure, functional domains, and subcellular localization of these LSR transcripts. Follow-up experiments will seek to verify these predictions in vitro and in vivo.
