Expression and Localization Profiles of Rhoptry Proteins in Plasmodium berghei Sporozoites.

Naohito Tokunaga,Takafumi Tsuboi,Mamoru Nozaki,Kazuhiro Matsuoka,Tomoko Ishino,Minami Baba,Motomi Torii,Mayumi Tachibana

doi:10.3389/fcimb.2019.00316

Naohito Tokunaga, Takafumi Tsuboi + Show 6 more

Open Access

https://doi.org/10.3389/fcimb.2019.00316

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Abstract

In the Plasmodium lifecycle two infectious stages of parasites, merozoites, and sporozoites, efficiently infect mammalian host cells, erythrocytes, and hepatocytes, respectively. The apical structure of merozoites and sporozoites contains rhoptry and microneme secretory organelles, which are conserved with other infective forms of apicomplexan parasites. During merozoite invasion of erythrocytes, some rhoptry proteins are secreted to form a tight junction between the parasite and target cell, while others are discharged to maintain subsequent infection inside the parasitophorous vacuole. It has been questioned whether the invasion mechanisms mediated by rhoptry proteins are also involved in sporozoite invasion of two distinct target cells, mosquito salivary glands and mammalian hepatocytes. Recently we demonstrated that rhoptry neck protein 2 (RON2), which is crucial for tight junction formation in merozoites, is also important for sporozoite invasion of both target cells. With the aim of comprehensively describing the mechanisms of sporozoite invasion, the expression and localization profiles of rhoptry proteins were investigated in Plasmodium berghei sporozoites. Of 12 genes representing merozoite rhoptry molecules, nine are transcribed in oocyst-derived sporozoites at a similar or higher level compared to those in blood-stage schizonts. Immuno-electron microscopy demonstrates that eight proteins, namely RON2, RON4, RON5, ASP/RON1, RALP1, RON3, RAP1, and RAMA, localize to rhoptries in sporozoites. It is noteworthy that most rhoptry neck proteins in merozoites are localized throughout rhoptries in sporozoites. This study demonstrates that most rhoptry proteins, except components of the high-molecular mass rhoptry protein complex, are commonly expressed in merozoites and sporozoites in Plasmodium spp., which suggests that components of the invasion mechanisms are basically conserved between infective forms independently of their target cells. Combined with sporozoite-stage specific gene silencing strategies, the contribution of rhoptry proteins in invasion mechanisms can be described.

Highlights

During the malaria lifecycle, three invasive forms, ookinetes, sporozoites, and merozoites, invade different types of cells in mosquito vectors and mammalian hosts
From the catalog of known P. falciparum rhoptry proteins (Counihan et al, 2013), 12 molecules whose orthologous genes exist in P. berghei were selected to compare their expression between merozoites and sporozoites
In this study it is demonstrated that most rhoptry proteins reported in merozoites are expressed in sporozoites, with the exception of components of the high-molecular mass rhoptry protein (RhopH) complex

Summary

Introduction

Three invasive forms, ookinetes, sporozoites, and merozoites, invade different types of cells in mosquito vectors and mammalian hosts. Sporozoites, which transmit malaria disease from mosquitoes to mammalian hosts, firstly invade mosquito salivary glands prior to be injected into the mammalian skin during a blood meal. Micronemes in the highly motile sporozoite and ookinete stages contain proteins involved in motility and cell traversal (Sultan et al, 1997; Yuda and Ishino, 2004; Kariu et al, 2006). Rhoptries are present only in infective stage parasites, such as Plasmodium merozoites and sporozoites which proliferate inside the PVM, and are absent in ookinetes, raising the possibility that rhoptry secretory proteins are involved in cell infection (reviewed in Baum et al, 2008; Frenal et al, 2017)

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