Gliding motility protein LIMP promotes optimal mosquito midgut traversal and infection by Plasmodium berghei

CTRP (circumsporozoite protein and thrombospondin-related adhesive protein [TRAP]-related protein) of the rodent malaria parasite Plasmodium berghei (PbCTRP) makes up a protein family together with other apicomplexan proteins that are specifically expressed in the host-invasive stage 1. PbCTRP is produced in the mosquito-invasive, or ookinete, stage and is a protein candidate for a role in ookinete adhesion and invasion of the mosquito midgut epithelium. To demonstrate involvement of PbCTRP in the infection of the vector, we performed targeting disruption experiments with this gene. PbCTRP disruptants showed normal exflagellation rates and development into ookinetes. However, no oocyst formation was observed in the midgut after ingestion of these parasites, suggesting complete loss of their invasion ability. On the other hand, when ingested together with wild-type parasites, disruptants were able to infect mosquitoes, indicating that the PbCTRP gene of the wild-type parasite rescued infectivity of disruptants when they heterologously mated in the mosquito midgut lumen. Our results show that PbCTRP plays a crucial role in malaria infection of the mosquito midgut and suggest that similar molecular mechanisms are used by malaria parasites to invade cells in the insect vector and the mammalian host.

Download Full-text

SSP3 Is a Novel Plasmodium yoelii Sporozoite Surface Protein with a Role in Gliding Motility

Infection and Immunity ◽

10.1128/iai.01800-14 ◽

2014 ◽

Vol 82 (11) ◽

pp. 4643-4653 ◽

Cited By ~ 18

Author(s):

Anke Harupa ◽

Brandon K. Sack ◽

Viswanathan Lakshmanan ◽

Nadia Arang ◽

Alyse N. Douglass ◽

...

Keyword(s):

Salivary Glands ◽

Biological Activities ◽

Surface Protein ◽

Plasmodium Yoelii ◽

Gliding Motility ◽

Surface Proteins ◽

Type I ◽

Content Type ◽

Mosquito Midgut

ABSTRACTPlasmodiumsporozoites develop within oocysts in the mosquito midgut wall and then migrate to the salivary glands. After transmission, they embark on a complex journey to the mammalian liver, where they infect hepatocytes. Proteins on the sporozoite surface likely mediate multiple steps of this journey, yet only a few sporozoite surface proteins have been described. Here, we characterize a novel, conserved sporozoite surface protein (SSP3) in the rodent malaria parasitePlasmodium yoelii. SSP3 is a putative type I transmembrane protein unique toPlasmodium. By using epitope tagging and SSP3-specific antibodies in conjunction with immunofluorescence microscopy, we showed that SSP3 is expressed in mosquito midgut oocyst sporozoites, exhibiting an intracellular localization. In sporozoites derived from the mosquito salivary glands, however, SSP3 localized predominantly to the sporozoite surface as determined by immunoelectron microscopy. However, the ectodomain of SSP3 appeared to be inaccessible to antibodies in nonpermeabilized salivary gland sporozoites. Antibody-induced shedding of the major surface protein circumsporozoite protein (CSP) exposed the SSP3 ectodomain to antibodies in some sporozoites. Targeted deletion ofSSP3adversely affectedin vitrosporozoite gliding motility, which, surprisingly, impacted neither their cell traversal capacity, host cell invasionin vitro, nor infectivityin vivo. Together, these data reveal a previously unappreciated complexity of thePlasmodiumsporozoite surface proteome and the roles of surface proteins in distinct biological activities of sporozoites.

Download Full-text

Zygote morphogenesis but not the establishment of cell polarity in Plasmodium berghei is controlled by the small GTPase, RAB11A

10.1101/811802 ◽

2019 ◽

Author(s):

H. Patil ◽

K.R. Hughes ◽

L. Lemgruber ◽

N. Philip ◽

N. Dickens ◽

...

Keyword(s):

Plasmodium Berghei ◽

Protein Transport ◽

Sexual Transmission ◽

Small Gtpase ◽

Mammalian Host ◽

Mosquito Vector ◽

Cellular Polarity ◽

Morphological Transformation ◽

Mosquito Midgut

AbstractPlasmodium species are apicomplexan parasites whose zoites are polarized cells with a marked apical organisation where the organelles associated with host cell invasion and colonization reside. Plasmodium gametes mate in the mosquito midgut to form the spherical and presumed apolar zygote that morphs during the following 24 hours into a polarized, elongated and motile zoite form, the ookinete. Endocytosis-mediated protein transport is generally necessary for the establishment and maintenance of polarity in epithelial cells and neurons, and the small GTPase RAB11A is an important regulator of protein transport via recycling endosomes. PbRAB11A is essential in blood stage asexual of Plasmodium. Therefore, a promoter swap strategy was employed to down-regulate PbRAB11A expression in gametocytes and zygotes of the rodent malaria parasite, Plasmodium berghei which demonstrated the essential role of RAB11A in ookinete development. The approach revealed that lack of PbRAB11A had no effect on gamete production and fertility rates however, the zygote to ookinete transition was almost totally inhibited and transmission through the mosquito was prevented. Lack of PbRAB11A did not prevent meiosis and mitosis, nor the establishment of polarity as indicated by the correct formation and positioning of the Inner Membrane Complex (IMC) and apical complex. However, morphological maturation was prevented and parasites remained spherical and immotile and furthermore, they were impaired in the secretion and distribution of microneme cargo. The data are consistent with the previously proposed model of RAB11A endosome mediated delivery of plasma membrane in Toxoplasma gondii if not its role in IMC formation and implicate it in microneme function.Author SummaryAccording to the WHO there was estimated to be over 200 million cases of malaria in 2017 and nearly half a million deaths. The disease is caused by specific species of Plasmodium which are passed between human hosts by a mosquito vector. In order to transmit through the mosquito the single-celled parasite undergoes many developmental changes as it morphs from non-motile blood forms to become a polarised and motile ookinete in the mosquito midgut. Transport of proteins within the cell during these critical morphological transitions relies on specific endosome vesicles to correctly target proteins within the parasite. We investigated the role of the RAB11A protein which is known to be involved in endosomal vesicle targeting to generate cellular polarity in other organisms. Because RAB11A is also essential for parasite growth in the mammalian host we used a promoter swap system to specifically switch off RAB11A in the sexual transmission stages. In the absence of RAB11A parasites were unable to form elongated, motile ookinetes and were unable to pass through the mosquito. Interestingly the parasites were able to form some of the (polarising) structures specific to ookinetes however full morphological transformation did not occur and the parasites were not motile. We show that although proteins are still delivered to the parasite surface, secretion is impaired and that the mutant parasites are smaller despite obvious microtubule formation implying that there is a deficit in delivery of membrane to the surface.

Download Full-text

A single-chain antibody fragment specific for the Plasmodium berghei ookinete protein Pbs21 confers transmission blockade in the mosquito midgut

Molecular and Biochemical Parasitology ◽

10.1016/s0166-6851(99)00158-9 ◽

1999 ◽

Vol 104 (2) ◽

pp. 195-204 ◽

Cited By ~ 37

Author(s):

Shigeto Yoshida ◽

Hiroyuki Matsuoka ◽

Enjie Luo ◽

Kuni Iwai ◽

Meiji Arai ◽

...

Keyword(s):

Plasmodium Berghei ◽

Antibody Fragment ◽

Single Chain ◽

Single Chain Antibody ◽

Mosquito Midgut ◽

Single Chain Antibody Fragment

Download Full-text

Conserved peptide sequences bind to actin and enolase on the surface of Plasmodium berghei ookinetes

Parasitology ◽

10.1017/s0031182011001296 ◽

2011 ◽

Vol 138 (11) ◽

pp. 1341-1353 ◽

Cited By ~ 15

Author(s):

J. HERNÁNDEZ-ROMANO ◽

M. H. RODRÍGUEZ ◽

V. PANDO ◽

J. A. TORRES-MONZÓN ◽

A. ALVARADO-DELGADO ◽

...

Keyword(s):

Electron Microscopy ◽

Phage Display ◽

Plasmodium Berghei ◽

Surface Proteins ◽

Peptide Sequence ◽

Specific Antibodies ◽

Mosquito Midgut ◽

Complex Process ◽

Consensus Peptide ◽

Immunofluorescence And Electron Microscopy

SUMMARYThe description of Plasmodium ookinete surface proteins and their participation in the complex process of mosquito midgut invasion is still incomplete. In this study, using phage display, a consensus peptide sequence (PWWP) was identified in phages that bound to the Plasmodium berghei ookinete surface and, in selected phages, bound to actin and enolase in overlay assays with ookinete protein extracts. Actin was localized on the surface of fresh live ookinetes by immunofluorescence and electron microscopy using specific antibodies. The overall results indicated that enolase and actin can be located on the surface of ookinetes, and suggest that they could participate in Plasmodium invasion of the mosquito midgut.

Download Full-text

Plasmodium berghei Sporozoites Acquire Virulence and Immunogenicity during Mosquito Hemocoel Transit

Infection and Immunity ◽

10.1128/iai.00758-13 ◽

2013 ◽

Vol 82 (3) ◽

pp. 1164-1172 ◽

Cited By ~ 24

Author(s):

Yuko Sato ◽

Georgina N. Montagna ◽

Kai Matuschewski

Keyword(s):

Salivary Glands ◽

Plasmodium Berghei ◽

Lower Proportion ◽

Gliding Motility ◽

Mammalian Host ◽

Live Attenuated Vaccine ◽

Experimental Cerebral Malaria ◽

Content Type ◽

Vector Borne

ABSTRACTMalaria is a vector-borne disease caused by the single-cell eukaryotePlasmodium. The infectious parasite forms are sporozoites, which originate from midgut-associated oocysts, where they eventually egress and reach the mosquito hemocoel. Sporozoites actively colonize the salivary glands in order to be transmitted to the mammalian host. Whether residence in the salivary glands provides distinct and vital cues for the development of infectivity remains unsolved. In this study, we systematically compared the infectivity ofPlasmodium bergheisporozoites isolated from the mosquito hemocoel and salivary glands. Hemocoel sporozoites display a lower proportion of gliding motility but develop into liver stages when added to cultured hepatoma cells or after intravenous injection into mice. Mice infected by hemocoel sporozoites had blood infections similar to those induced by sporozoites liberated from salivary glands. These infected mice display indistinguishable systemic inflammatory cytokine responses and develop experimental cerebral malaria. When used as metabolically active, live attenuated vaccine, hemocoel sporozoites elicit substantial protection against sporozoite challenge infections. Collectively, these findings show that salivary gland colonization does not influence parasite virulence in the mammalian host when sporozoites are administered intravenously. This conclusion has important implications forin vitrosporozoite production and manufacturing of whole-sporozoite vaccines.

Download Full-text

Role of Plasmodium berghei ookinete surface and oocyst capsule protein, a novel oocyst capsule-associated protein, in ookinete motility

10.21203/rs.3.rs-326425/v1 ◽

2021 ◽

Author(s):

Kazuhiko Nakayama ◽

Yuta Kimura ◽

Yu Kitahara ◽

Akira Soga ◽

Asako Haraguchi ◽

...

Keyword(s):

Blood Meal ◽

Plasmodium Berghei ◽

Protein A ◽

Gliding Motility ◽

Wild Type ◽

Oocyst Wall ◽

Infected Blood ◽

Capsule Formation

Abstract Background: Plasmodium sp., which causes malaria, must first develop in mosquitoes before being transmitted. Upon ingesting infected blood, gametes form in the mosquito lumen, followed by fertilization and differentiation of the resulting zygotes into motile ookinetes. Within 24 hours of blood ingestion, these ookinetes traverse mosquito epithelial cells and lodge below the midgut basal lamina, where they differentiate into sessile oocysts that are protected by a capsule. Methods: We identified an ookinete surface and oocyst capsule protein (OSCP) that is involved in ookinete motility as well as oocyst capsule formation. Results: We found that knockout of OSCP in parasite decreases ookinete gliding motility and gradually reduces the number of oocysts. On day 15 after blood ingestion, the oocyst wall was significantly thinner. Moreover, adding anti-OSCP antibodies decreased the gliding speed of wild type ookinetes in vitro. Adding anti-OSCP antibodies to an infected blood meal also resulted in decreased oocyst formation. Conclusion: These findings may be useful for the development of transmission-blocking tools for malaria.

Download Full-text

Structure of an atypical homodimeric actin capping protein from the malaria parasite

10.1101/2020.08.16.253187 ◽

2020 ◽

Author(s):

Ábris Ádám Bendes ◽

Petri Kursula ◽

Inari Kursula

Keyword(s):

Crystal Structure ◽

Actin Filament ◽

Plasmodium Berghei ◽

Malaria Parasite ◽

Gliding Motility ◽

Actin Dynamics ◽

Structural Elements ◽

Β Subunit ◽

Capping Proteins ◽

Actin Capping

AbstractActin capping proteins (CPs) are essential regulators of actin dynamics in all eukaryotes. Their structure and function have been extensively characterized in higher eukaryotes but their role and mechanism of action in apicomplexan parasites remain enigmatic. Here, we present a crystal structure of a unique homodimeric CP from the rodent malaria parasite Plasmodium berghei. In addition, we compare homo- and heterodimeric arrangements of P. berghei CPs (PbCPs) in solution. We complement our findings by describing the regulatory effects of PbCPs on heterologous skeletal muscle α-actin as well as parasite actin. Comprehensive kinetic and steadystate measurements show atypical regulation of actin dynamics; PbCPs facilitate rapid turnover of parasite actin I without affecting the apparent critical concentration. Possibly to rescue actin filament capping in life cycle stages where the CP β-subunit is downregulated, homo- and heterodimeric PbCPs show redundant effects in vitro. However, our data suggest that homodimers may in addition influence actin kinetics by recruiting lateral actin dimers. This unusual function could arise from the absence of a β-subunit, as the asymmetric PbCP homodimer lacks the structural elements essential for canonical barbed end interactions, suggesting a novel CP binding mode. These findings facilitate further studies aimed at elucidating the precise actin filament capping mechanism in Plasmodium and the eligibility of PbCPs as drug targets against malaria.Significance statementMalaria parasites of the genus Plasmodium display a unique form of gliding motility, which depends on an unconventional actomyosin motor. Actin capping proteins (CPs) play a major role in regulating parasite motility. Here, we describe a unique Plasmodium berghei CP (PbCP) system, behaving contradictory to canonical heterodimeric CPs, more suited to regulate the fast dynamics of the parasite actin system. We present the crystal structure of a distinctive homodimeric form of PbCP and extensive biochemical data, describing the atypical behavior of each PbCP form. The PbCP homodimer displays capping even in the absence of canonical conserved structural elements, suggesting a novel actin-CP interaction mode. These distinct structural properties could provide opportunities for drug design against malaria.

Download Full-text