Structural Insights Into PfARO and Characterization of its Interaction With PfAIP

Geiger, Michael; Brown, Chris; Wichers, Jan Stephan; Strauss, Jan; Lill, Andrés; Thuenauer, Roland; Liffner, Benjamin; Wilcke, Louisa; Lemcke, Sarah; Heincke, Dorothee; Pažický, Samuel; Bachmann, Anna; Löw, Christian; Wilson, Danny W.; Filarsky, Michael; Burda, Paul‐Christian; Zhang, Kun; Junop, M.S.; Gilberger, Tim‐Wolf

doi:10.1016/j.jmb.2019.12.024

Cited by 24 publications

(32 citation statements)

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“…It is likely that rhoptry and PPM fusion are controlled by proteins that localise to the cytosolic face of the rhoptry membrane, as they can contact both membranes to facilitate their fusion. To date, however, only three proteins in P. falciparum have been shown to localise to the cytosolic face of the rhoptry membrane: A rmadillo r epeats o nly (ARO) 7 , A RO i nteracting p rotein (AIP) 8 , and C ytosolically e xposed r hoptry l eaflet i nteracting protein 1 (CERLI1; also known as R hoptry a pical s urface p rotein (RASP) 2) 9,10 . Here we describe PfCERLI2 (Pf3D7_0405200), a paralogue of Pfcerli1 , which localises to the cytosolic face of the rhoptry bulb membrane and is essential for merozoite invasion of RBCs.…”

Section: Introductionmentioning

confidence: 99%

Pfcerli2, a duplicated gene in the malaria parasitePlasmodium falciparumessential for invasion of erythrocytes as revealed by phylogenetic and cell biological analysis

Liffner

Balbin

Shami

et al. 2020

Preprint

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Merozoite invasion of host red blood cells (RBCs) is essential for survival of the human malaria parasite Plasmodium falciparum. Proteins involved with RBC binding and invasion are secreted from dual-club shaped organelles at the apical tip of the merozoite called the rhoptries. Here we characterise P. falciparum Cytosolically Exposed Rhoptry Leaflet Interacting protein 2 (PfCERLI2), as a rhoptry bulb protein that is essential for merozoite invasion. Phylogenetic analyses show that cerli2 arose through an ancestral gene duplication of cerli1, a related cytosolically exposed rhoptry bulb protein. We show that PfCERLI2 is essential for blood-stage growth and localises to the cytosolic face of the rhoptry bulb. Inducible knockdown of PfCERLI2 led to an inhibition of merozoite invasion after tight junction formation. PfCERLI2 knockdown was associated with inhibition of rhoptry antigen processing and a significant elongation of the rhoptries, suggesting that the inability of merozoites to invade is caused by aberrant rhoptry function due to PfCERLI2 deficiency. These findings identify PfCERLI2 as a protein that has key roles in rhoptry biology during merozoite invasion.

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Section: Introductionmentioning

confidence: 99%

Pfcerli2, a duplicated gene in the malaria parasitePlasmodium falciparumessential for invasion of erythrocytes as revealed by phylogenetic and cell biological analysis

Liffner

Balbin

Shami

et al. 2020

Preprint

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“…Approximately 30 proteins have been linked to the rhoptry 8 . Despite the rhoptries being membrane-bound organelles with dynamic functions during secretion of invasion ligands, there remains only a single protein reported to localise to the luminal face (Rhoptry associated membrane antigen 12 ) and two to the cytosolic face (Armadillo repeats only 13 , ARO Interacting Protein 14,15 ) of the P. falciparum rhoptry membrane. Therefore, the drivers of rhoptry secretion and compartmentalisation are unknown.…”

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confidence: 99%

PfCERLI1 is a conserved rhoptry associated protein essential for Plasmodium falciparum merozoite invasion of erythrocytes

et al. 2020

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The disease-causing blood-stage of the Plasmodium falciparum lifecycle begins with invasion of human erythrocytes by merozoites. Many vaccine candidates with key roles in binding to the erythrocyte surface and entry are secreted from the large bulb-like rhoptry organelles at the apical tip of the merozoite. Here we identify an essential role for the conserved protein P. falciparum Cytosolically Exposed Rhoptry Leaflet Interacting protein 1 (PfCERLI1) in rhoptry function. We show that PfCERLI1 localises to the cytosolic face of the rhoptry bulb membrane and knockdown of PfCERLI1 inhibits merozoite invasion. While schizogony and merozoite organelle biogenesis appear normal, biochemical techniques and semi-quantitative superresolution microscopy show that PfCERLI1 knockdown prevents secretion of key rhoptry antigens that coordinate merozoite invasion. PfCERLI1 is a rhoptry associated protein identified to have a direct role in function of this essential merozoite invasion organelle, which has broader implications for understanding apicomplexan invasion biology.

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“…In order to exclude a localization with the apical organelles we co-localized PfAAP6-GFP and PfAAP4-GFP with appropriate marker proteins: PfARO (Cabrera et al, 2012;Geiger et al, 2020) (rhoptry bulb marker), PfRON12 (Knuepfer et al, 2014;Ito et al, 2019) or PfAIP (Geiger et al, 2020) (rhoptry neck marker) and PfAMA1 (Peterson et al, 1989;Healer et al, 2002) (micronemes marker) and additionally PfAAP4-GFP with…”

Section: Distinct Localization Of Apical Annuli Protein Homologues Inmentioning

confidence: 99%

“…PfKELCH13 (Birnbaum et al, 2020) and PfAIP (Geiger et al, 2020). To achieve mislocalization, we used our PIC4-2xFKBP-GFP-2xFKBP parasites line and transfected it with the 1xNLS-FRB-mCherry plasmid (Birnbaum et al, 2017) encoding the nuclear mislocalizer coupled to FRB.…”

Section: Functional Assessment Of Pic1 Pic2 Pic3 Pic4 Pfaap4 Pfamentioning

confidence: 99%

“…For co-localization experiments, the full length sequence of organellar marker genes for micronemes (ama1 Pf3D7_1133400) (Peterson et al, 1989;Healer et al, 2002), rhoptry neck (aip, Pf3D7_1136700; ron12 Pf3D7_1017100) (Knuepfer et al, 2014;Ito et al, 2019;Geiger et al, 2020), inner membrane complex (imc1c/alv5, Pf3D7_1003600) (Hu et al, 2010) were amplified from 3D7 cDNA and for dense granules (exp2, PF3D7_1471100) (Bullen et al, 2012) from the plasmid pEXP2rec (Mesén-Ramírez et al, 2019) and cloned into pARL-ama1-ARO-mCherry-BSD (Cabrera et al, 2012) using the AvrII/KpnI restriction site or in pARL-ama1-ARO-mCherry-yDHODH (derived from pARL-ama1-ARO-mCherry-BSD (Cabrera et al, 2012)).…”

Section: Cloning Of Dna Constructsmentioning

confidence: 99%

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Identification of novel inner membrane complex and apical annuli proteins of the malaria parasitePlasmodium falciparum

Wichers

Wunderlich

Heincke

et al. 2021

Preprint

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The inner membrane complex (IMC) is a defining feature of apicomplexan parasites, which confers stability and shape to the cell, functions as a scaffolding compartment during the formation of daughter cells and plays an important role in motility and invasion during different life cycle stages of these single celled organisms. To explore the IMC proteome of the malaria parasite Plasmodium falciparum we applied a proximity-dependent biotin identification (BioID)-based proteomics approach, using the established IMC marker protein Photosensitized INA-Labelled protein 1 (PhIL1) as bait in asexual blood-stage parasites. Subsequent mass spectrometry-based peptide identification revealed enrichment of twelve known IMC proteins and several uncharacterized candidate proteins. We validated nine of these previously uncharacterized proteins by endogenous GFP-tagging. Six of these represent new IMC proteins, while three proteins have a distinct apical localization that most likely represent structures described as apical annuli in Toxoplasma gondii. Additionally, various Kelch13 interacting candidates were identified, suggesting an association of the Kelch13 compartment and the IMC in schizont and merozoite stages. This work extends the number of validated IMC proteins in the malaria parasite and reveals for the first time the existence of apical annuli proteins in P. falciparum. Additionally, it provides evidence for a spatial association between the Kelch13 compartment and the IMC in late blood-stage parasites.

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Structural Insights Into PfARO and Characterization of its Interaction With PfAIP

Cited by 24 publications

References 61 publications

Pfcerli2, a duplicated gene in the malaria parasitePlasmodium falciparumessential for invasion of erythrocytes as revealed by phylogenetic and cell biological analysis

Pfcerli2, a duplicated gene in the malaria parasitePlasmodium falciparumessential for invasion of erythrocytes as revealed by phylogenetic and cell biological analysis

PfCERLI1 is a conserved rhoptry associated protein essential for Plasmodium falciparum merozoite invasion of erythrocytes

Identification of novel inner membrane complex and apical annuli proteins of the malaria parasitePlasmodium falciparum

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