Discipline: Biological Sciences
Subcategory: Cell and Molecular Biology
Room: Exhibit Hall
Alexandra LaFollette - Boise State University
Co-Author(s): Dr. Troy Rohn, Boise State University, Boise, IDDr. Julia Oxford, Boise State University, Boise, IDSaylor Leising, Boise State University, Boise, IDMadyson McCarthy, Boise State University, Boise, ID
Alzheimer’s is a fatal disorder that causes neuronal cell loss in brain regions crucial for thinking and memory. While the inheritance risk of the APOE4 allele has been well documented, the molecular basis for how ApoE4 leads to enhanced dementia risk is not yet understood. Previous work in the Rohn lab has shown that proteolysis and formation of an amino-terminal fragment (ApoE4 1-151) may contribute to toxicity in vitro. Using zebrafish as an in vivo model organism, mutants expressing this fragment were generated to examine any deleterious effects. As an initial approach, this study seeks to determine the lethality of these transgenic F1 zebrafish. We hypothesize that expression of the ApoE4 (1-151) amino-terminal fragment in transgenic zebrafish will lead to an increased mortality when compared to wild-type (WT) zebrafish. To assess toxicity of this fragment in F1 mutant zebrafish, 30 mutant zebrafish and 30 WT zebrafish will be analyzed. Once the larvae are at 72 hours post fertilization, they will be screened for the expression of this protein fragment using GFP and RFP. The two groups of fish will then be observed for a period of 45 days post hatching until they reach the juvenile developmental stage. During the 45 days, survivorship rates will be recorded. A total of three trials will be performed, and at the end of each trial the data will be analyzed using the Kaplan-Meier probability test and the log-rank method to determine significance. For trial one, there were 9/30 WT deaths and 16/30 mutant deaths. The probability of survival for WT was 70% and for mutants it was 46.7%. Data analysis returned a p-value of 0.080, lacking overall significance. Although the hypothesis has initially been disproved, future trials may help support the idea that the ApoE4 fragment is lethal in F1 mutants. Additionally, it is hypothesized that the fragment will be more lethal in homozygous F2 generations. Thus, more trials of the survivorship study need to be performed to accurately assess how fatal possessing the ApoE4 (1-151) fragment is. Data from this study will help to illuminate the toxicity of this amino terminal protein fragment in vivo and how it may contribute to the pathology of Alzheimer’s disease. In the future, homozygous F2 generation mutants will be bred once F1 mutants reach an appropriate age. After the mutant lines have been propagated, tissue will be collected from both WT and mutant zebrafish. The tissue will be fixed, sliced, and stained to characterize differences in morphology as well as localization of the ApoE4 (1-151) intracellularly by confocal analysis. Future experiments will also be conducted on the mutant strain and compared to the WT strain to assess memory and motor deficits.
Funder Acknowledgement(s): The Bridges to Baccalaureate program is supported by the National Institute of General Medical Sciences of the National Institutes of Health under Award No. R25GM123927. This project was further supported by the Center of Excellence in Biomedical Research through the Institutional Development Award (IDeA) from the National Institute of General Medical Sciences of the National Institutes of Health under Grant Nos. P20GM109095 and P20GM103408 and the National Science Foundation S-STEM Gateway Scholarships in Biological Sciences under Grant Award No. DUE-1644233. Additional support was received from the National Institutes of Health Grant No. 2R15AG042781-02A1 and the KO Alzheimer's Dementia Foundation in Boise, ID. We also acknowledge support from an Institutional Development Award (IDeA) from the National Institute of General Medical Sciences of the National Institutes of Health under Grant Nos. P20GM103408 and P20GM109095.
Faculty Advisor: Dr. Troy Rohn, email@example.com
Role: The part of the research I have done was basically taking this project from start to finish. Aside from creating the transgenic fish and raising them to breeding age, I have done all other breeding of the F0 line and have propagated these F1 mutants myself. I have also been the one to track their survivorship rates. In addition, I am part of the paper that proves toxicity of this fragment in vitro by performing experiments with the full length ApoE4 protein vs the 1-151 fragment. I will also play a role in the future work listed in this abstract once the fish reach an appropriate age.