Glutathione Reductase-null Malaria Parasites Have Normal Blood Stage Growth but Arrest during Development in the Mosquito

Malaria parasites contain a complete glutathione (GSH) redox system, and several enzymes of this system are considered potential targets for antimalarial drugs. Through generation of a gamma-glutamylcysteine synthetase (gamma-GCS)-null mutant of the rodent parasite Plasmodium berghei, we previously showed that de novo GSH synthesis is not critical for blood stage multiplication but is essential for oocyst development. In this study, phenotype analyses of mutant parasites lacking expression of glutathione reductase (GR) confirmed that GSH metabolism is critical for the mosquito oocyst stage. Similar to what was found for gamma-GCS, GR is not essential for blood stage growth. GR-null parasites showed the same sensitivity to methylene blue and eosin B as wild type parasites, demonstrating that these compounds target molecules other than GR in Plasmodium. Attempts to generate parasites lacking both GR and gamma-GCS by simultaneous disruption of gr and gamma-gcs were unsuccessful. This demonstrates that the maintenance of total GSH levels required for blood stage survival is dependent on either de novo GSH synthesis or glutathione disulfide (GSSG) reduction by Plasmodium GR. Our studies provide new insights into the role of the GSH system in malaria parasites with implications for the development of drugs targeting GSH metabolism.