A cytoplasmic requirement of red cells for invasion by malarial parasites

Abstract

Human red cells, when lysed by dialysis at high haematocrit against a medium of low ionic strength and then dialysed back to physiological saline at 37 degrees C, give rise to resealed ghosts that are invaded with high efficiency by Plasmodium falciparum parasites. When the haematocrit is reduced, a critical concentration is reached, such that the resealed ghosts no longer support invasion. This indicates that a constituent of the cytoplasm becomes diluted to a concentration below a critical level. This constituent is evidently ATP, for when extraneous ATP is added to the diluent and the dialysate, the susceptibility to invasion is fully restored. This does not occur when the non-hydrolysable analogue, adenylyl-imidodiphosphate (AMP-PNP) is substituted for ATP, whereas the hydrolysable ATP analogue, adenosine-5'-O-(3-thiotriphosphate) (ATP-gamma-S), which can be utilised by kinases, can partly replace ATP. Stimulation of invasion by the addition of 2,3-diphosphoglycerate was also associated with a perceptible rise in ATP concentration. The invasion process does not appear to involve intracellular calcium, for EGTA introduced into the resealed ghost has no detectable effect. Moreover, vanadate in the medium does not appreciably inhibit invasion, and it is thus unlikely that the requirement for ATP is linked to the activity of membrane ion-pump enzymes. An inhibitor of phosphorylation, adenosine, introduced into the cells at high concentration, causes significant inhibition of invasion. The results suggest that ATP is required for maintaining the turnover of phosphoryl groups of membrane-associated proteins, such as spectrin. A basic scheme for the mechanism of the invasion process is suggested. In addition to the effect of ATP, it is also shown that with greater dilution, and in the presence of ATP, there is an abrupt loss of susceptibility to invasion. It is inferred that this is due to the dilution of another essential cytoplasmic constituent to below a critical concentration. This second constituent has not yet been identified.