Malaria caused by the parasite Plasmodium falciparum is responsible for approximately half a million deaths annually (WHO, 2017). The development of antimalarial resistance from P. falciparum elicits an urgent need to understand parasite biology and develop novel therapeutics.

This summer, I had the opportunity to continue working with Dr. Audrey John at the Children’s Hospital of Philadelphia. Her lab looks to understand the metabolic pathways in the parasite as a means to gain insight on potential novel drug targets. Under her mentorship, I researched the impact of loss of the HAD2 enzyme on drug resistance and parasite growth.

My project builds off previous lab members’ work to determine the role of the HAD2 enzyme in maintaining metabolic homeostasis and exhibiting resistance to the drug fosmidomycin (FSM). FSM is a competitive inhibitor of the DXR enzyme, which the parasite needs to synthesize isoprenoids, a class of biomolecules involved in essential cellular functions such as respiration. Our lab found that one strain of FSM-resistant parasites possessed a single nucleotide polymorphism in the gene encoding HAD2 that resulted in a truncated protein. In addition to being FSM-resistant, this strain, had2R157X, was growth-attenuated relative to the wild-type. Therefore, we hypothesized that HAD2 is required to maintain homeostasis between glycolysis and isoprenoid biosynthesis.

To test this, I used CRISPR/Cas9 genome editing techniques to transfect a wild-type parental strain with an inducible knockdown construct. This allows for “turning on” and “off” genes, and then assessing and characterizing the impact of near total depletion of HAD2 on the parasite. I ran assays to determine the half maximal inhibitory concentration of FSM against these strains when had2 was turned “on” and compared it to when it was “off,” verified the system worked by western blotting, and performed growth assays to see how HAD2 (or lack of it) impacts parasite growth.

Participating in this research project allowed me to apply the knowledge I’ve learned in my STEM courses to a project with real implications, which was incredibly satisfying. More than just reading about western blots in my genetics and molecular biology classes, I was able to actually run them myself, from sample prep to imaging. This gave me a greater appreciation for all the work gone into research projects and a greater understanding of the importance of science. Seeing this project through the summer, I learned how to ask important questions, how to design hypothesis-driven experiments, and how to format data to show the main takeaway effectively. Beyond the day-to-day work, participating in the research project also strengthened my resolve to pursue a career in research. Thank you to the Frances Velay Fellowship and the Ruth M. Kanter College Alumni Undergraduate Research Grant for making this experience possible!