Characterization of the Tubovesicular Network in Plasmodium vivax Liver Stage Hypnozoites and Schizonts

Plasmodium is a genus of apicomplexan parasites which replicate in the liver before causing malaria. Plasmodium vivax can also persist in the liver as dormant hypnozoites and cause clinical relapse upon activation, but the molecular mechanisms leading to activation have yet to be discovered. In this study, we use high-resolution microscopy to characterize temporal changes of the P. vivax liver stage tubovesicular network (TVN), a parasitophorous vacuole membrane (PVM)-derived network within the host cytosol. We observe extended membrane clusters, tubules, and TVN-derived vesicles present throughout P. vivax liver stage development. Additionally, we demonstrate an unexpected presence of the TVN in hypnozoites and observe some association of this network to host nuclei. We also reveal that the host water and solute channel aquaporin-3 (AQP3) associates with TVN-derived vesicles and extended membrane clusters. AQP3 has been previously shown to localize to the PVM of P. vivax hypnozoites and liver schizonts but has not yet been shown in association to the TVN. Our results highlight host-parasite interactions occur in both dormant and replicating liver stage P. vivax forms and implicate AQP3 function during this time. Together, these findings enhance our understanding of P. vivax liver stage biology through characterization of the TVN with an emphasis on the presence of this network in dormant hypnozoites.

Download Full-text

Plasmodium berghei EXP-1 interacts with host Apolipoprotein H during Plasmodium liver-stage development

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1606419114 ◽

2017 ◽

Vol 114 (7) ◽

pp. E1138-E1147 ◽

Cited By ~ 22

Author(s):

Cláudia Sá e Cunha ◽

Britta Nyboer ◽

Kirsten Heiss ◽

Margarida Sanches-Vaz ◽

Diana Fontinha ◽

...

Keyword(s):

Parasitophorous Vacuole ◽

Malaria Prevention ◽

Rapid Expansion ◽

Parasite Protein ◽

Liver Stage ◽

Apolipoprotein H ◽

Replication Phase ◽

Parasite Infections ◽

Stage Development ◽

Host Parasite

The first, obligatory replication phase of malaria parasite infections is characterized by rapid expansion and differentiation of single parasites in liver cells, resulting in the formation and release of thousands of invasive merozoites into the bloodstream. Hepatic Plasmodium development occurs inside a specialized membranous compartment termed the parasitophorous vacuole (PV). Here, we show that, during the parasite’s hepatic replication, the C-terminal region of the parasitic PV membrane protein exported protein 1 (EXP-1) binds to host Apolipoprotein H (ApoH) and that this molecular interaction plays a pivotal role for successful Plasmodium liver-stage development. Expression of a truncated EXP-1 protein, missing the specific ApoH interaction site, or down-regulation of ApoH expression in either hepatic cells or mouse livers by RNA interference resulted in impaired intrahepatic development. Furthermore, infection of mice with sporozoites expressing a truncated version of EXP-1 resulted in both a significant reduction of liver burden and delayed blood-stage patency, leading to a disease outcome different from that generally induced by infection with wild-type parasites. This study identifies a host–parasite protein interaction during the hepatic stage of infection by Plasmodium parasites. The identification of such vital interactions may hold potential toward the development of novel malaria prevention strategies.

Download Full-text

The tubovesicular network in Plasmodium vivax liver-stage hypnozoites and schizonts associates with host aquaporin 3

10.1101/2020.12.09.417949 ◽

2020 ◽

Author(s):

Kayla Sylvester ◽

Steven P. Maher ◽

Dora Posfai ◽

Michael K. Tran ◽

McKenna C. Crawford ◽

...

Keyword(s):

High Resolution ◽

Parasitophorous Vacuole ◽

Parasitophorous Vacuole Membrane ◽

Liver Stage ◽

Vacuole Membrane ◽

Aquaporin 3 ◽

Host Parasite Interactions ◽

Host Parasite ◽

High Resolution Microscopy ◽

Stage Infection

AbstractThe apicomplexan Plasmodium parasites replicate in the liver before causing malaria. P. vivax can also persist in the liver as dormant hypnozoites and cause relapses upon activation. The host water and solute channel aquaporin-3 (AQP3) has been shown to localize to the parasitophorous vacuole membrane (PVM) of P. vivax hypnozoites and liver schizonts, along with other Plasmodium species and stages. In this study, we use high-resolution microscopy to characterize temporal changes of the tubovesicular network (TVN), a PVM-derived network within the host cytosol, during P. vivax liver-stage infection. We demonstrate an unexpected role for the TVN in hypnozoites and reveal AQP3 associates with TVN-derived vesicles and extended membrane features. We further show AQP3 recruitment to Toxoplasma gondii. Our results highlight dynamic host-parasite interactions that occur in both dormant and replicating liver-stage P. vivax forms and implicate AQP3 function during this time. Together, these findings enhance our understanding of AQP3 in apicomplexan infection.

Download Full-text

Plasmodium vivax Liver Stage Development and Hypnozoite Persistence in Human Liver-Chimeric Mice

Cell Host & Microbe ◽

10.1016/j.chom.2015.03.015 ◽

2015 ◽

Vol 17 (4) ◽

pp. 536 ◽

Cited By ~ 1

Author(s):

Sebastian A. Mikolajczak ◽

Ashley M. Vaughan ◽

Niwat Kangwanrangsan ◽

Wanlapa Roobsoong ◽

Matthew Fishbaugher ◽

...

Keyword(s):

Plasmodium Vivax ◽

Human Liver ◽

Chimeric Mice ◽

Liver Stage ◽

Stage Development

Download Full-text

Immunologic Characterization of Plasmodium Vivax Antigens Using Plasmodium Cynomolgi Liver Stage-Primed Immune Sera

American Journal of Tropical Medicine and Hygiene ◽

10.4269/ajtmh.1994.51.365 ◽

1994 ◽

Vol 51 (3) ◽

pp. 365-371 ◽

Cited By ~ 4

Author(s):

Chunfu Yang ◽

William E. Collins ◽

Pascal Millet

Keyword(s):

Plasmodium Vivax ◽

Liver Stage ◽

Plasmodium Cynomolgi

Download Full-text

Discovery of New Microneme Proteins in Cryptosporidium parvum and Implication of the Roles of a Rhomboid Membrane Protein (CpROM1) in Host–Parasite Interaction

Frontiers in Veterinary Science ◽

10.3389/fvets.2021.778560 ◽

2021 ◽

Vol 8 ◽

Author(s):

Xin Gao ◽

Jigang Yin ◽

Dongqiang Wang ◽

Xiaohui Li ◽

Ying Zhang ◽

...

Keyword(s):

Parasitophorous Vacuole ◽

Immunofluorescence Assay ◽

Host Cells ◽

Immunogold Electron Microscopy ◽

Apicomplexan Parasites ◽

Subsequent Transformation ◽

Dense Granules ◽

Veterinary Importance ◽

Host Parasite ◽

Microneme Proteins

Apicomplexan parasites possess several unique secretory organelles, including rhoptries, micronemes, and dense granules, which play critical roles in the invasion of host cells. The molecular content of these organelles and their biological roles have been well-studied in Toxoplasma and Plasmodium, but are underappreciated in Cryptosporidium, which contains many parasites of medical and veterinary importance. Only four proteins have previously been identified or proposed to be located in micronemes, one of which, GP900, was confirmed using immunogold electron microscopy (IEM) to be present in the micronemes of intracellular merozoites. Here, we report on the discovery of four new microneme proteins (MICs) in the sporozoites of the zoonotic species C. parvum, identified using immunofluorescence assay (IFA). These proteins are encoded by cgd3_980, cgd1_3550, cgd1_3680, and cgd2_1590. The presence of the protein encoded by cgd3_980 in sporozoite micronemes was further confirmed using IEM. Cgd3_980 encodes one of the three C. parvum rhomboid peptidases (ROMs) and is, thus, designated CpROM1. IEM also confirmed the presence of CpROM1 in the micronemes of intracellular merozoites, parasitophorous vacuole membranes (PVM), and feeder organelles (FO). CpROM1 was enriched in the pellicles and concentrated at the host cell–parasite interface during the invasion of sporozoites and its subsequent transformation into trophozoites. CpROM1 transcript levels were also higher in oocysts and excysted sporozoites than in the intracellular parasite stages. These observations indicate that CpROM1, an intramembrane peptidase with membrane proteolytic activity, is involved in host–parasite interactions, including invasion and proteostasis of PVM and FO.

Download Full-text

Hijacking of the host cell Golgi by Plasmodium liver stage parasites

10.1101/2020.07.24.220137 ◽

2020 ◽

Author(s):

Mariana De Niz ◽

Gesine Kaiser ◽

Benoit Zuber ◽

Won Do Heo ◽

Volker T. Heussler ◽

...

Keyword(s):

Host Cell ◽

Parasitophorous Vacuole ◽

Dominant Negative ◽

Rab Gtpases ◽

Cell Organelles ◽

Liver Stage ◽

Golgi Fragmentation ◽

Stage Development ◽

Golgi Structure ◽

Dynamic Fluorescence

AbstractThe intracellular lifestyle represents a challenge for the rapidly proliferating liver stage Plasmodium parasite. In order to scavenge host resources, Plasmodium has evolved the ability to target and manipulate host cell organelles. Using dynamic fluorescence-based imaging, we show a direct interplay between the pre-erythrocytic stages of Plasmodium berghei and the host cell Golgi during the entire liver stage development. Liver stage schizonts fragment the host cell Golgi into miniaturized stacks, which increases surface interactions with the parasite’s parasitophorous vacuole membrane. Interference with the host cell Golgi-linked vesicular machinery using specific dominant-negative Arf and Rab GTPases results in developmental arrest and diminished survival of liver stage parasites. Moreover, functional Rab11a is critical for the parasites ability to induce Golgi fragmentation. Altogether, we demonstrate that the structural and functional integrity of the host cell Golgi is necessary for optimal pre-erythrocytic development of P. berghei. The parasite hijacks the hepatocyte’s Golgi structure to optimize its own intracellular development.

Download Full-text

Transcriptome and histone epigenome of Plasmodium vivax salivary-gland sporozoites point to tight regulatory control and potential mechanisms for liver-stage differentiation

10.1101/145250 ◽

2017 ◽

Cited By ~ 1

Author(s):

◽

Ivo Muller ◽

Aaron R. Jex ◽

Stefan H. I. Kappe ◽

Sebastian A. Mikolajczak ◽

...

Keyword(s):

Salivary Gland ◽

Plasmodium Vivax ◽

Translational Control ◽

Regulatory Control ◽

Small Subset ◽

Liver Stage ◽

Mixed Blood ◽

Differential Transcription ◽

Liver Infection

ABSTRACTPlasmodium vivax is the key obstacle to malaria elimination in Asia and Latin America, largely attributed to its ability to form resilient hypnozoites (sleeper-cells) in the host liver that escape treatment and cause relapsing infections. The decision to form hypnozoites is made early in the liver infection and may already be set in sporozoites prior to invasion. To better understand these early stages of infection, we undertook a comprehensive transcriptomic and histone epigenetic characterization of P. vivax sporozoites. The salivary-gland sporozoite transcriptome is heavily composed of transcripts associated with functions needed for early infection of the vertebrate host and development within hepatocytes. Through comparisons to recently published proteome data for the P. vivax sporozoite, our study finds that although highly transcribed, these transcripts are not detectable as proteins and may be regulated through translational repression; a finding we test for a small subset of transcripts and proteins through immunofluorescent microscopy of sporozoites and liver stages in humanized mice. We identify differential transcription between the sporozoite and published transcriptomes of asexual blood-stages and mixed versus hypnozoite-enriched liver stages. These comparisons point to multiple layers of transcriptional, post-transcriptional and post-translational control that appear active in sporozoites and to a lesser extent hypnozoites, but largely absent in replicating liver schizonts or mixed blood-stages. Common transcripts up-regulated in sporozoites and hypnozoites compared to mixed (i.e., schizont) liver-stages identify genes linked to dormancy/persistence in bacteria, amoebae and plants. We also characterise histone epigenetic modifications in the P. vivax sporozoite and explore their role in regulating transcription. Collectively, these data support the hypothesis that the sporozoite as a tightly programmed stage primed to infect the human host and identifies potential mechanisms for hypnozoite-formation that may be further explored in liver stage models.

Download Full-text

CRISPR/Cas9-Based Knockout of GNAQ Reveals Differences in Host Cell Signaling Necessary for Egress of Apicomplexan Parasites

mSphere ◽

10.1128/msphere.01001-20 ◽

2020 ◽

Vol 5 (6) ◽

pp. e01001-20

Author(s):

Paul-Christian Burda ◽

Hugo Bisio ◽

Jean-Baptiste Marq ◽

Dominique Soldati-Favre ◽

Volker T. Heussler

Keyword(s):

Host Cell ◽

Parasitophorous Vacuole ◽

Host Cells ◽

Infected Host ◽

Protein Subunit ◽

Host Cell Membrane ◽

Guanine Nucleotide Binding Protein ◽

Liver Stage ◽

Content Type ◽

Apicomplexan Parasites

ABSTRACTToxoplasma gondii and members of the genus Plasmodium are obligate intracellular parasites that leave their infected host cell upon a tightly controlled process of egress. Intracellular replication of the parasites occurs within a parasitophorous vacuole, and its membrane as well as the host plasma membrane need to be disrupted during egress, leading to host cell lysis. While several parasite-derived factors governing egress have been identified, much less is known about host cell factors involved in this process. Previously, RNA interference (RNAi)-based knockdown and antibody-mediated depletion identified a host signaling cascade dependent on guanine nucleotide-binding protein subunit alpha q (GNAQ) to be required for the egress of Toxoplasma tachyzoites and Plasmodium blood stage merozoites. Here, we used CRISPR/Cas9 technology to generate HeLa cells deficient in GNAQ and tested their capacity to support the egress of T. gondii tachyzoites and Plasmodium berghei liver stage parasites. While we were able to confirm the importance of GNAQ for the egress of T. gondii, we found that the egress of P. berghei liver stages was unaffected in the absence of GNAQ. These results may reflect differences between the lytic egress process in apicomplexans and the formation of host cell-derived vesicles termed merosomes by P. berghei liver stages.IMPORTANCE The coordinated release of apicomplexan parasites from infected host cells prior to reinvasion is a critical process for parasite survival and the spread of infection. While Toxoplasma tachyzoites and Plasmodium blood stages induce a fast disruption of their surrounding membranes during their egress from host cells, Plasmodium liver stages keep the host cell membrane intact and leave their host cell in host cell-derived vesicles called merosomes. The knockout of GNAQ, a protein involved in G-protein-coupled receptor signaling, demonstrates the importance of this host factor for the lytic egress of T. gondii tachyzoites. Contrastingly, the egress of P. berghei is independent of GNAQ at the liver stage, indicating the existence of a mechanistically distinct strategy to exit the host cell.

Download Full-text

Plasmodium falciparum PF10_0164 (ETRAMP10.3) Is an Essential Parasitophorous Vacuole and Exported Protein in Blood Stages

Eukaryotic Cell ◽

10.1128/ec.00336-09 ◽

2010 ◽

Vol 9 (5) ◽

pp. 784-794 ◽

Cited By ~ 23

Author(s):

Drew C. MacKellar ◽

Matthew T. O'Neill ◽

Ahmed S. I. Aly ◽

John B. Sacci ◽

Alan F. Cowman ◽

...

Keyword(s):

Plasmodium Falciparum ◽

Parasitophorous Vacuole ◽

Structural Similarity ◽

Plasmodium Yoelii ◽

Parasitophorous Vacuole Membrane ◽

Liver Stage ◽

Content Type ◽

Vacuole Membrane ◽

Stage Development ◽

Gene 4

ABSTRACT Upregulated in infectious sporozoites gene 4 (UIS4) encodes a parasitophorous vacuole membrane protein expressed in the sporozoite and liver stages of rodent malaria parasites. Parasites that lack UIS4 arrest in early liver-stage development, and vaccination of mice with uis4 − sporozoites confers sterile protection against challenge with infectious sporozoites. Currently, it remains unclear whether an ortholog of UIS4 is carried in the human malaria parasite Plasmodium falciparum, although the gene PF10_0164 has been identified as a candidate ortholog for UIS4 on the basis of synteny and structural similarity of the encoded protein. We show that PF10_0164 is expressed in sporozoites and blood stages of P. falciparum, where it localizes to the parasitophorous vacuole, and is also exported to the host erythrocyte. PF10_0164 is refractory to disruption in asexual blood stages. Functional complementation was tested in Plasmodium yoelii by replacing the endogenous copy of UIS4 with PF10_0164. PF10_0164 localized to the parasitophorous vacuole membrane of liver stages, but transgenic parasites did not complete liver-stage development in mice. We conclude that PF10_0164 is a parasitophorous vacuole protein that is essential in asexual blood stages and that does not complement P. yoelii UIS4, and it is thus likely not a functional ortholog of UIS4.

Download Full-text