The overarching goal of my research is to use a variety of model organisms to elucidate the basic mechanisms which orchestrate the fantastic assembly of a growing embryo. Understanding these mechanisms will lay the foundation for biomedical research aimed at treating and preventing disorders at both ends of our human existence, birth and age-related death.
My PhD thesis research is being performed in the lab of Dr. Maria Garcia-Garcia. Pictures coming!
We used ethylnitrosourea (ENU), a chemical mutagen, to generate mutations in the germ line of mice and then examined the offspring of these mice for embryonic birth defects. The goal was to find embryos which exhibited defects in early organ formation in order to identify novel mechanisms which govern these early steps in development. My research in the lab is focused on two classes of mutants which possess a brain defect and heart defect, respectively. The focus of this ongoing research is to identify the underlying mutation which is causing each defect and then determine which mechanisms are interrupted when these genes are mutated.
Figure 1 - Genetic factors which impact gut tilting.
During my transition into the PhD program at Cornell University, I took the opportunity to get exposure to other model systems through my rotation research. I spent one rotation in the lab of Dr. Natasza Kurpios studying the mechanisms of left-right asymmetry during gut rotation in chicken embryos. As it elongates, the gut begins to bend toward the left, a carefully coordinated movement controlled by structural shape changes in the supporting dorsal mesentery (Figure 1). Previously, the Kurpios lab had compared gene expression levels between the left and right dorsal mesentery and identified genes which were enriched on one side or the other. I was able to verify through in situ hybridizations that several components of Notch signaling were enriched on the left side of the dorsal mesentery. Though currently unknown, we proposed that these Notch components conditioned the left dorsal mesentery to allow blood vessels to pass from the body to the gut tube. I also spent a rotation in the lab of Dr. Adrienne Roeder where I studied the process of waxy cuticle formation on Arabidopsis sepals. We used scanning electron microscopy to study mutants which possessed cuticle defects. This, along with live imaging of newly forming sepals demonstrated that the cuticle develops as a wavefront moving from apical to basal on the growing sepal.
I had the privilege of working in the lab of Dr. Hui-Min Chung at the University of West Florida. Dr. Chung's lab works with Drosophila melanogaster and, in 2011 incorporated red flour beetles (Tribolium castaneum) into some undergraduate projects. I've worked along-side many great students during those years; the individuals shown below do not represent the current lab members, but rather, the individuals I worked with:
Joel BrownGrad (2012)
Joanna BridgesGrad (2013)
Anthony PintoVisiting Grad (2010)
Liz KennedyVisiting Grad (2011)
Erica PoundsUndergrad (2011)
Nick SpencerUndergrad (2011)
Thomas StephensonUndergrad (2011)
Rainey BoothUndergrad (2012)
Preston ShisgalUndergrad (2012)
Meghan Kirby Undergrad (2012)
Figure 2 - Gamma-Secretase Complex (Parks. 2007)
We used the fruit fly, Drosophila melanogaster, as a genetic model to study the function of gamma-secretase. Gamma-secretase is a protease which plays a critical role in Notch signaling, a cellular signaling mechanism which operates during the development of every major body system. In humans, ?-secretase also cleaves Amyloid Precursor Protein (APP), resulting in fragments called beta-amyloid peptides which readily aggregate to form the beta-amyloid plaques present in Alzheimer's Disease (AD) patients.
Gamma-secretase is composed of four subunits: Aph-1, Nicastrin, Presenilin, and Pen-2 (Figure 2). While Presenilin is the active component of this protease, it depends on the other components for assembly (Figure 3). It was previously shown that in the absence of Pen-2, the ?-Secretase complex failed to assemble. I was interested in determining whether, in addition to its role in assembly, Pen-2 was also required for the function of the complex. In order to examine this, I used fly development to assess how the absence of Pen-2 would affect Notch signaling. As expected, in the absence of Pen-2, Notch signaling did not operate. However, we also showed that by replacing the normal Presenilin protein with an altered Presenilin protein, Notch signaling persisted even in the absence of Pen-2. Together these results suggest that while Pen-2 is not required for the function of ?-secretase, it does act to stabilize the normal mature ?-secretase complex. A better understanding of how ?-secretase functions will aid in understanding how it promotes neurodegeneration in Alzheimer’s disease patients.
Figure 3 - Assembly of Gamma-Secretase Complex (Iwatsubo. 2004)
Barakat, A., Mercer, B., Cooper, E., Chung, H., 2009. Examining Requirement for Formation of Functional Presenilin Proteins and their Processing Events in Vivo. Genesis The Journal of Genetics and Development 3.
Cooper, E., Deng, W., Chung, H., 2009. Aph-1 is Required to Regulate Presenilin-Mediated Gamma-Secretase Activity and Cell Survival in Drosophila Wing Development. Genesis The Journal of Genetics and Development 3.
Iwatsubo, T. (2004). The ?-secretase complex: Machinery for intramembrane proteolysis. Current Opinion in Neurobiology, 14(3), 379-383. doi:DOI: 10.1016/j.conb.2004.05.010
Parks, A. L., & Curtis, D. (2007). Presenilin diversifies its portfolio. Trends in Genetics, 23(3), 140-150. doi:DOI: 10.1016/j.tig.2007.01.008