Perforin‐like protein PPLP 2 permeabilizes the red blood cell membrane during egress of P lasmodium falciparum gametocytes

Abstract

Egress of malaria parasites from the host cell requires the concerted rupture of its enveloping membranes. Hence, we investigated the role of the plasmodial perforin-like protein PPLP2 in the egress of Plasmodium falciparum from erythrocytes. PPLP2 is expressed in blood stage schizonts and mature gametocytes. The protein localizes in vesicular structures, which in activated gametocytes discharge PPLP2 in a calcium-dependent manner. PPLP2 comprises a MACPF domain and recombinant PPLP2 has haemolytic activities towards erythrocytes. PPLP2-deficient [PPLP2(−)] merozoites show normal egress dynamics during the erythrocytic replication cycle, but activated PPLP2(−) gametocytes were unable to leave erythrocytes and stayed trapped within these cells. While the parasitophorous vacuole membrane ruptured normally, the activated PPLP2(−) gametocytes were unable to permeabilize the erythrocyte membrane and to release the erythrocyte cytoplasm. In consequence, transmission of PPLP2(−) parasites to the Anopheles vector was reduced. Pore-forming equinatoxin II rescued both PPLP2(−) gametocyte exflagellation and parasite transmission. The pore sealant Tetronic 90R4, on the other hand, caused trapping of activated wild-type gametocytes within the enveloping erythrocytes, thus mimicking the PPLP2(−) loss-of-function phenotype. We propose that the haemolytic activity of PPLP2 is essential for gametocyte egress due to permeabilization of the erythrocyte membrane and depletion of the erythrocyte cytoplasm.

The development of malaria parasites in the mosquito midgut

“…These egress vesicles contain the plasmodial pore‐forming perforin PPLP2, which is able to perforate the EM. This event occurs approximately 6 min post‐activation of the gametocyte and results in the release of the erythrocyte cytoplasm; hence, the gamete is then contained by the EM only (Deligianni et al ., 2013; Wirth et al ., 2014). Approximately 15 min after uptake by the blood‐feeding mosquito, the EM opens and releases the fertile gamete (Sologub et al ., 2011; Wirth et al ., 2014; reviewed in Wirth and Pradel, 2012).…”

Phospholipases during membrane dynamics in malaria parasites

“…Also, while the PbPL-deficient parasites undergo replication and develop merozoites, these are unable to egress from the host hepatocyte (Burda et al, 2015). In this context, perforin-like proteins have previously been identified and shown to be important for parasite egress from the HC in a calciumdependent manner (Deligianni et al, 2013;Garg et al, 2013;Wirth et al, 2014). It is thus suggested that during egress PbPL acts in a conglomerate of different phospholipases or pore-forming proteins.…”

In silico analysis of calcium binding pocket of perforin like protein 1: insights into the regulation of pore formation

“…plays a pivotal role in the physiology of malaria parasite Garg et al 2013;Singh et al 2010). It regulates invasion and egress of malaria parasites through various molecular players, perforin like proteins (PLPs) being one of them Ecker et al 2007;Garg et al 2013;Ishino et al 2005;Kaiser et al 2004;Kadota et al 2004;Singh et al 2010;Wirth et al 2014). These proteins help in transversal of malaria parasite from one cell to the other in liver and mosquito stages (Ecker et al 2007;Ishino et al 2005;Kadota et al 2004).…”

“…These proteins help in transversal of malaria parasite from one cell to the other in liver and mosquito stages (Ecker et al 2007;Ishino et al 2005;Kadota et al 2004). Recently, they are shown to play a crucial role in the egress of malaria parasites in the blood stage and gametocyte stages Wirth et al 2014). Being proteins of multistage functional relevance, these are usually an interest of the researchers to develop them as a target for intervention of parasite growth at various stages of life cycle.…”

“…Plasmodium falciparum PLPs (PfPLPs) are multi domain proteins with an N-terminal signal sequence, a central MACPF domain and a C-terminal domain (Garg et Wirth et al 2014). The lytic activity of these PfPLPs is due to a structurally conserved, pore forming central MACPF domain.…”