GRA47 is important for the morphology and permeability of the parasitophorous vacuole in Toxoplasma gondii

Establishing an intact intracellular parasitophorous vacuole (PV) that enables efficient nutrient uptake and protein trafficking is essential for the survival and proliferation of Toxoplasma gondii. Although the PV membrane (PVM)-localized dense granule protein 17 (GRA17) and GRA23 mediate the permeability of the PVM to small molecules, including nutrient uptake and excretion of metabolic by-products, the molecular mechanism by which T. gondii acquires nutrients remains unclear. In this study, we showed that the secreted protein GRA47 contributed to normal PV morphology, PVM permeability to small molecules, growth, and virulence in T. gondii. Co-immunoprecipitation analysis demonstrated potential interaction of GRA47 with GRA72, and the loss of GRA72 affected PV morphology, parasite growth and infectivity. To investigate the biological relationship among GRA47, GRA72, GRA17 and GRA23, attempts were made to construct strains with double gene deletion and overexpressing strains. Only Delta gra23 Delta gra72 was successfully constructed. This strain exhibited a significant increase in the proportion of aberrant PVs compared with the Delta gra23 strain. Overexpressing one of the three related GRAs partially rescued PVs with aberrant morphology in Delta gra47, Delta gra72 and Delta gra17, while the expression of the Plasmodium falciparum PVM protein PfExp2, an ortholog of GRA17 and GRA23, fully rescued the PV morphological defect in all three Delta gra strains. These results suggest that these GRA proteins may not be functionally redundant but rather work in different ways to regulate nutrient acquisition. These findings highlight the versatility of the nutrient uptake mechanisms in T. gondii, which may contribute to the parasite's remarkable ability to grow in different cellular niches in a very broad range of hosts. (c) 2024 Australian Society for Parasitology. Published by Elsevier Ltd. All rights are reserved, including those for text and data mining, AI training, and similar technologies.