Establishing Acanthamoeba as a modern model parasite for infectious disease biology and comparative metabolism by defining mitochondrial proteins and pathways

PROJECT SUMMARY / ABSTRACT This proposal presents a five-year research and career development plan focused on establishing Acanthamoeba castellanii—the third leading cause of infectious blindness and a cause of fatal brain infections—as a modern model parasite for studying infectious disease biology, comparative metabolism, and mitochondrial molecular biology, specifically by defining its mitochondrial proteins and pathways. The candidate, Jonathan Stefely, MD, PhD, completed clinical pathology training at Massachusetts General Hospital (MGH) and is now a postdoctoral researcher under the mentorship of Dr. Vamsi Mootha at MGH. Dr. Mootha is a pioneer in mitochondrial biology, genetics, and biochemistry, and has trained 80 scientists, including 29 postdoctoral scientists that now hold independent scientific leadership positions. The proposed project builds on Dr. Stefely’s previous research experience in biochemistry, now extending this work into unstudied areas of parasitology through an innovative and robust collaboration with local expert parasitology advisors, facilitated by his primary mentor, Dr. Mootha. Protozoan parasites span diverse branches of eukaryotic biology and contain strikingly diverse mitochondria, which are a proven anti-parasite drug target. Yet, most parasite mitochondrial biochemistry remains undefined. A central thesis of this proposal is that a systematic characterization of unique Acanthamoeba mitochondrial biochemistry that is not conserved in humans will unlock both new biology and new drug targets. In preliminary work with a team of collaborators at MGH, the Broad Institute, and Boston University, Dr. Stefely completely re-annotated the nuclear genome of Acanthamoeba castellanii and generated a high-quality draft of a new mitochondrial proteome inventory. Here, Dr. Stefely proposes to finalize and validate this Acanthamoeba “MitoCarta” (Aim 1) and to characterize select parasite mitochondrial proteins and pathways that were uncovered through his work, starting with an atypical branched electron transport chain (Aim 2) and novel proteins that function on mitochondrial DNA (mtDNA) (Aim 3). These three independent aims will build new knowledge of mitochondria, electron transport chains, mtDNA molecular biology, and potential anti-parasite drug targets. This mentored career development project will provide new training and expertise in parasitology, mitochondrial physiology, anoxic biology, and molecular biology—robustly supported by an exceptional group of mentors, advisors, collaborators, and unique resources at MGH, the Broad Institute, and Boston University. This project will also enable Dr. Stefely’s transition to independence with a focus on using the new Acanthamoeba genome annotation and MitoCarta to turn Acanthamoeba into a first-class model parasite for providing fundamental insights into the biology of protozoan parasites in clinical pathology.