Project Grant R01AI184855

PROTON PUMPING AND ENERGY METABOLISM OF THE RING STAGE MALARIA PARASITES - MALARIA REMAINS A MAJOR GLOBAL HEALTH BURDEN, AFFECTING 40% OF THE WORLD'S POPULATION. IN 2021, THERE WERE 247 MILLION REPORTED CASES, RESULTING IN OVER 619,000 DEATHS. THE DISEASE IS CAUSED BY REPETITIVE GROWTH OF THE PROTOZOAN PARASITES KNOWN AS PLASMODIUM SPP. INSIDE RED BLOOD CELLS (RBCS), LEADING TO RBC LYSIS. AMONG THE FIVE MALARIA SPECIES INFECTING HUMANS, PLASMODIUM FALCIPARUM IS THE MOST LETHAL, RESPONSIBLE FOR MORE THAN 80% OF THE DISEASE'S MORBIDITY AND MORTALITY. THIS PARASITE UNDERGOES ASEXUAL REPRODUCTION WITHIN RBCS OVER ITS 48-HOUR LIFECYCLE, WHICH CONSISTS OF THREE MAJOR STAGES: THE RING STAGE (~ 20 HOURS), THE TROPHOZOITE STAGE (~ 16 HOURS) AND THE SCHIZONT STAGE (~12 HOURS). DURING THE RING STAGE, INFECTED RBCS (IRBCS) HAVE A SMOOTH SURFACE LIKE UNINFECTED RBCS, ENABLING THEM TO EVADE CLEARANCE BY THE SPLEEN AND CIRCULATE IN THE PERIPHERAL BLOODSTREAM. THIS STAGE IS METABOLICALLY QUIESCENT AND CHALLENGING TO TARGET WITH AVAILABLE ANTIMALARIALS, AS MOST OF THESE DRUGS ARE NOT HIGHLY EFFECTIVE AGAINST THE SLOW-GROWING PARASITE. FURTHERMORE, THE RING STAGE CAN ENTER A DORMANT STATE AFTER DRUG TREATMENT, CONTRIBUTING TO DRUG RESISTANCE. THEREFORE, THERE IS AN URGENT NEED FOR A BETTER UNDERSTANDING OF THE FUNDAMENTAL BIOLOGY OF RING STAGE MALARIA PARASITES. OUR LABORATORY HAS MADE A SIGNIFICANT DISCOVERY BY REVEALING THAT THE METABOLICALLY INACTIVE RING STAGE PARASITE EMPLOYS AN ANCIENT PYROPHOSPHATE-DRIVEN PROTON PUMP TO SURVIVE FOR 20 HOURS INSIDE THE RBC FOLLOWING THE TIME OF INVASION. THIS ATP-INDEPENDENT PROTON PUMP IS KNOWN AS PFVP1 (PLASMODIUM FALCIPARUM VACUOLAR PYROPHOSPHATASE 1), AND IT DERIVES ENERGY FROM THE HYDROLYSIS OF PYROPHOSPHATE (PPI), A METABOLIC BY-PRODUCT GENERATED DURING SYNTHESIS OF DNA, RNA, AND PROTEIN. BASED ON THESE OBSERVATIONS, WE HYPOTHESIZE THAT THE RING-STAGE P. FALCIPARUM RELIES ON AN ATP-INDEPENDENT MECHANISM FOR PROTON EXPORTING AND ESTABLISHING PLASMA MEMBRANE POTENTIAL, IDENTIFYING A DISTINCTIVE VULNERABILITY IN THIS CHALLENGING STAGE OF THE ASEXUAL LIFECYCLE. TO TEST THIS HYPOTHESIS, WE OUTLINE TWO SPECIFIC AIMS. AIM I. INVESTIGATE THE IMPACT OF PFVP1'S LOOP SEQUENCES ON ITS PROTON PUMPING FUNCTIONALITY. AIM II. EXAMINE POLYPHOSPHATE METABOLISM AND PPI METABOLISM TO UNDERSTAND THE ENERGY SOURCE FOR PFVP1. THE OUTCOMES OF THIS RESEARCH PROPOSAL WILL PROVIDE INSIGHTS INTO HOW THE METABOLICALLY QUIESCENT RING STAGE PARASITE SURVIVES WITHIN THE RBC FOR NEARLY A DAY FOLLOWING INVASION. GIVEN THE ESSENTIAL NATURE OF PFVP1 AND ITS ABSENCE IN HUMANS, PFVP1 EMERGES AS AN IDEAL TARGET FOR COMBATING THE RING STAGE PARASITES. INHIBITING THIS LESS ACTIVE STAGE IS OF UTMOST IMPORTANCE TO ADVANCE MALARIA CONTROL AND ERADICATION EFFORTS.