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Rounding precedes rupture and breakdown of vacuolar membranes minutes before malaria parasite egress from erythrocytes.

ABSTRACT: Because Plasmodium falciparum replicates inside of a parasitophorous vacuole (PV) within a human erythrocyte, parasite egress requires the rupture of two limiting membranes. Parasite Ca2+ , kinases, and proteases contribute to efficient egress; their coordination in space and time is not known. Here, the kinetics of parasite egress were linked to specific steps with specific compartment markers, using live-cell microscopy of parasites expressing PV-targeted fluorescent proteins, and specific egress inhibitors. Several minutes before egress, under control of parasite [Ca2+ ]i , the PV began rounding. Then after ~1.5 min, under control of PfPKG and SUB1, there was abrupt rupture of the PV membrane and release of vacuolar contents. Over the next ~6 min, there was progressive vacuolar membrane deterioration simultaneous with erythrocyte membrane distortion, lasting until the final minute of the egress programme when newly formed parasites mobilised and erythrocyte membranes permeabilised and then ruptured-a dramatic finale to the parasite cycle of replication.

SUBMITTER: Glushakova S

PROVIDER: S-EPMC6368063 | biostudies-literature | 2018 Oct

REPOSITORIES: biostudies-literature

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Rounding precedes rupture and breakdown of vacuolar membranes minutes before malaria parasite egress from erythrocytes.

Glushakova Svetlana S Beck Josh R JR Garten Matthias M Busse Brad L BL Nasamu Armiyaw S AS Tenkova-Heuser Tatyana T Heuser John J Goldberg Daniel E DE Zimmerberg Joshua J

Cellular microbiology 20180710 10

Because Plasmodium falciparum replicates inside of a parasitophorous vacuole (PV) within a human erythrocyte, parasite egress requires the rupture of two limiting membranes. Parasite Ca<sup>2+</sup> , kinases, and proteases contribute to efficient egress; their coordination in space and time is not known. Here, the kinetics of parasite egress were linked to specific steps with specific compartment markers, using live-cell microscopy of parasites expressing PV-targeted fluorescent proteins, and s ...[more]

PMID: 29900649

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Project description:BackgroundEgress of Plasmodium falciparum, from erythrocytes at the end of its asexual cycle and subsequent parasite invasion into new host cells, is responsible for parasite dissemination in the human body. The egress pathway is emerging as a coordinated multistep programme that extends in time for tens of minutes, ending with rapid parasite extrusion from erythrocytes. While the Ca2+ regulation of the invasion of P. falciparum in erythrocytes is well established, the role of Ca2+ in parasite egress is poorly understood. This study analysed the involvement of cytoplasmic free Ca2+ in infected erythrocytes during the multistep egress programme of malaria parasites.MethodsLive-cell fluorescence microscopy was used to image parasite egress from infected erythrocytes, assessing the effect of drugs modulating Ca2+ homeostasis on the egress programme.ResultsA steady increase in cytoplasmic free Ca2+ is found to precede parasite egress. This increase is independent of extracellular Ca2+ for at least the last two hours of the cycle, but is dependent upon Ca2+ release from internal stores. Intracellular BAPTA chelation of Ca2+ within the last 45 minutes of the cycle inhibits egress prior to parasitophorous vacuole swelling and erythrocyte membrane poration, two characteristic morphological transformations preceding parasite egress. Inhibitors of the parasite endoplasmic reticulum (ER) Ca2+-ATPase accelerate parasite egress, indicating that Ca2+ stores within the ER are sufficient in supporting egress. Markedly accelerated egress of apparently viable parasites was achieved in mature schizonts using Ca2+ ionophore A23187. Ionophore treatment overcomes the BAPTA-induced block of parasite egress, confirming that free Ca2+ is essential in egress initiation. Ionophore treatment of immature schizonts had an adverse effect inducing parasitophorous vacuole swelling and killing the parasites within the host cell.ConclusionsThe parasite egress programme requires intracellular free Ca2+ for egress initiation, vacuole swelling, and host cell cytoskeleton digestion. The evidence that parasitophorous vacuole swelling, a stage of unaffected egress, is dependent upon a rise in intracellular Ca2+ suggests a mechanism for ionophore-inducible egress and a new target for Ca2+ in the programme liberating parasites from the host cell. A regulatory pathway for egress that depends upon increases in intracellular free Ca2+ is proposed.

Dataset Information

Rounding precedes rupture and breakdown of vacuolar membranes minutes before malaria parasite egress from erythrocytes.

Publications

Rounding precedes rupture and breakdown of vacuolar membranes minutes before malaria parasite egress from erythrocytes.

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