Melatonin and IP3-induced Ca2+ Release from Intracellular Stores in the Malaria Parasite Plasmodium falciparum within Infected Red Blood Cells

Eduardo Alves,Paula J Bartlett,Celia R.S Garcia,Andrew P Thomas

doi:10.1074/jbc.m110.188474

Abstract

IP(3)-dependent Ca(2+) signaling controls a myriad of cellular processes in higher eukaryotes and similar signaling pathways are evolutionarily conserved in Plasmodium, the intracellular parasite that causes malaria. We have reported that isolated, permeabilized Plasmodium chabaudi, releases Ca(2+) upon addition of exogenous IP(3). In the present study, we investigated whether the IP(3) signaling pathway operates in intact Plasmodium falciparum, the major disease-causing human malaria parasite. P. falciparum-infected red blood cells (RBCs) in the trophozoite stage were simultaneously loaded with the Ca(2+) indicator Fluo-4/AM and caged-IP(3). Photolytic release of IP(3) elicited a transient Ca(2+) increase in the cytosol of the intact parasite within the RBC. The intracellular Ca(2+) pools of the parasite were selectively discharged, using thapsigargin to deplete endoplasmic reticulum (ER) Ca(2+) and the antimalarial chloroquine to deplete Ca(2+) from acidocalcisomes. These data show that the ER is the major IP(3)-sensitive Ca(2+) store. Previous work has shown that the human host hormone melatonin regulates P. falciparum cell cycle via a Ca(2+)-dependent pathway. In the present study, we demonstrate that melatonin increases inositol-polyphosphate production in intact intraerythrocytic parasite. Moreover, the Ca(2+) responses to melatonin and uncaging of IP(3) were mutually exclusive in infected RBCs. Taken together these data provide evidence that melatonin activates PLC to generate IP(3) and open ER-localized IP(3)-sensitive Ca(2+) channels in P. falciparum. This receptor signaling pathway is likely to be involved in the regulation and synchronization of parasite cell cycle progression.

Highlights

Cell (RBC)4 stage of Plasmodium life cycle is a clear pharmaceutical target
IP3 and DAG increases have been reported during P. falciparum gametocyte exflagellation involved in the sexual cycle and transmission to the mosquito vector [38] and Elabbadi et al, [36] reported ionomycin-induced elevations in IP3 in the asexual red blood cells (RBCs) stage of the life cycle, indicating an enzyme capable of PIP2 hydrolysis is present in P. falciparum
UV flash photolysis of caged-IP3 under these conditions elicited a rapid and transient increase in intracellular Ca2ϩ in RBCs infected with P. falciparum (Fig. 1A, representative trace of 81 cells from 15 independent experiments and Fig. 1B confocal images with Ca2ϩ changes shown in pseudocolor)

Summary

Introduction

Cell (RBC)4 stage of Plasmodium life cycle is a clear pharmaceutical target. The RBC cycle of P. falciparum occurs over a period of 48 h (the life cycles of other Plasmodium species are multiples of 24 h) and consists of three stages of parasite development known as ring, trophozoite, and schizont. These molecules were able to induce Ca2ϩ release from cultured P. falciparum and P. chabaudi and importantly these responses were blocked by PLC inhibition and melatonin receptor antagonism [14]. Photorelease of Caged IP3 Induces Ca2ϩ Mobilization in Intact P. falciparum—In this study, IP3-dependent Ca2ϩ release has been examined in intact P. falciparum within the host erythrocyte using flash photolysis of cell permeant caged-IP3.

Results

Conclusion