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Release of protein by erythrocytic stages of Plasmodium knowlesi during cultivation in vitro
Abstract:
Erythrocytes infected with schizonts of P. knowlesi and labelled in vitro with (3)H-isoleucine were shown to release radioactivity when placed in unlabelled culture medium containing uninfected erythrocytes. Release of radiolabel probably occurred during schizont rupture and invasion of new erythrocytes and was not observed to any great extent at earlier stages in the erythrocytic life cycle. Approximately 55% of the released radioactivity was soluble in 10% trichloroacetic acid, the remainder being acid insoluble and therefore presumably protein in nature. Particulate or aggregated material appeared to comprise 40-50% of the acid-insoluble radiolabelled fraction. Two major labelled polypeptide species were observed on sodium dodecyl sulfate polyacrylamide gel electrophoresis of samples of concentrated culture medium. These proteins had apparent molecular weights of 45 000 and 49 000, which correspond closely with the sizes of two main protein species previously identified in P. knowlesi-infected cells.
Insights
Plasmodium knowlesi infected erythrocytes release radiolabeled proteins during schizont rupture. These released proteins, including two major polypeptides, are crucial for infecting new erythrocytes.
Area of Science:
- Malariology
- Molecular Parasitology
- Cell Biology
Background:
- Plasmodium knowlesi is a significant cause of malaria in humans.
- Understanding parasite release mechanisms is vital for malaria control.
- Erythrocytic stages of Plasmodium parasites are responsible for disease pathology.
Purpose of the Study:
- To investigate the release of radiolabeled molecules during the erythrocytic cycle of Plasmodium knowlesi.
- To characterize the nature and molecular weight of released proteins.
- To determine the stage of the parasite life cycle associated with radiolabel release.
Main Methods:
- In vitro labeling of Plasmodium knowlesi infected erythrocytes with (3H)-isoleucine.
- Culturing labeled infected erythrocytes with uninfected erythrocytes in unlabeled medium.
- Analysis of released radioactivity using trichloroacetic acid precipitation.
- Characterization of acid-insoluble fractions via SDS-PAGE.
Main Results:
- Radiolabel release was observed during schizont rupture and erythrocyte invasion.
- Approximately 55% of released radioactivity was soluble in TCA, while the remainder was acid-insoluble (protein).
- Two major radiolabeled polypeptide species (45,000 and 49,000 Da) were identified in the culture medium, matching known Plasmodium knowlesi proteins.
Conclusions:
- Plasmodium knowlesi releases significant amounts of protein during the final stages of its erythrocytic cycle.
- These released proteins likely play a role in merozoite invasion of new erythrocytes.
- The identified proteins represent key parasite components involved in host cell invasion.