Biology subjects

Burda, P.-C.

Publications and source records attributed to Burda, P.-C..

10 recordsLinked to original sources

bioRxiv2026

A new class of lipid transfer proteins is required for the recycling of lipids from the P. falciparum digestive vacuole

Malaria parasites endocytose large quantities of hemoglobin from the host erythrocyte, a process critical for parasite survival, leading to extensive membrane internalization. While hemoglobin degradation in the digestive vacuole (DV) is well studied, how the parasite deals with the membranes arriving within the DV is unknown. Here we identified PfTUPA, a previously uncharacterized lipid transfer protein in the DV membrane that is needed for this function. PfTUPA contains a soluble TULIP-like lipid transport domain exposed to the DV lumen and a transmembrane lipid transfer domain of bacterial origin (PqiA) in the DV membrane. Structural comparisons revealed proteins with various PqiA and TULIP-like domain combinations across distant eukaryotic clades, indicating this is a frequent functional partnership. Hence, PfTUPA belongs to a new class of eukaryotic lipid transfer proteins that in malaria parasites is needed for a key function of its biology.

microbiology↗

bioRxiv2026

Plasmodium berghei liver stages establish a damage-mimicking vacuole to hijack host ER membrane contact site machinery for lipid uptake

Plasmodium liver stage parasites undergo massive intracellular replication and therefore require extensive acquisition of host-derived lipids. How parasites coordinate lipid supply at the host-parasite interface, however, remains poorly understood. Here, we show that the Plasmodium berghei parasitophorous vacuole membrane (PVM) selectively mimics features of damaged endolysosomal membranes, including dynamic LC3-positive membrane tubules and PI4K2A-dependent PI(4)P accumulation, while lacking canonical damage markers including ESCRT components and galectins. This damage-mimicking membrane state drives OSBP recruitment and is associated with formation of VAPA/B-positive ER-PVM membrane contact sites. Disruption of VAPA/B or OSBP impaired PVM integrity and parasite survival, while OSBP depletion reduced cholesterol accumulation at the PVM. Together, our findings suggest that liver stage parasites exploit host lysosome repair-like pathways to redirect host lipids toward parasite growth and intracellular survival. These results redefine the PVM as an active metabolic interface that enables host lipid acquisition rather than merely serving as a protective barrier.

microbiology↗

bioRxiv2023

Disruption of a Plasmodium falciparum patatin-like phospholipase delays male gametocyte exflagellation

An essential process in transmission of the malaria parasite to the Anopheles vector is the conversion of mature gametocytes into gametes within the mosquito gut, where they egress from the red blood cell (RBC). During egress, male gametocytes undergo exflagellation, leading to the formation of eight haploid motile microgametes, while female gametes retain their spherical shape. Gametocyte egress depends on sequential disruption of the parasitophorous vacuole membrane and the host cell membrane. In other life cycle stages of the malaria parasite, phospholipases have been implicated in membrane disruption processes during egress, however their importance for gametocyte egress is relatively unknown. Here, we performed comprehensive functional analyses of six putative phospholipases for their role during development and egress of Plasmodium falciparum gametocytes. We localize two of them, the prodrug activation and resistance esterase (PF3D7_0709700) and the lysophospholipase 1 (PF3D7_1476700), to the parasite plasma membrane. Subsequently, we show that disruption of most of the studied phospholipase genes does neither affect gametocyte development nor egress. The exception is the putative patatin-like phospholipase PF3D7_0924000, whose gene deletion leads to a delay in male gametocyte exflagellation, indicating an important, albeit not essential, role of this enzyme in male gametogenesis.

microbiology↗

bioRxiv2023

A malaria parasite phospholipase facilitates efficient asexual blood stage egress

Malaria parasite release (egress) from host red blood cells involves parasite-mediated membrane poration and rupture, thought to involve membrane-lytic effector molecules such as perforin-like proteins and/or phospholipases. With the aim of identifying these effectors, we disrupted the expression of two Plasmodium falciparum perforin-like proteins simultaneously and showed that they have no essential roles during blood stage egress. Proteomic profiling of parasite proteins discharged into the parasitophorous vacuole (PV) just prior to egress detected the presence in the PV of a lecithin:cholesterol acyltransferase (LCAT; PF3D7_0629300). Conditional ablation of LCAT resulted in abnormal egress and a reduced replication rate. Lipidomic profiles showed drastic changes in several phosphatidylserine and acylphosphatidylglycerol species during egress. We thus show that, in addition to its previously demonstrated role in liver stage merozoite egress, LCAT is required to facilitate efficient egress in asexual blood stage malaria parasites. Author SummaryMalaria kills over half a million people every year worldwide. It is caused by a single-celled parasite called Plasmodium falciparum that grows and multiplies within a bounding vacuole, inside red blood cells of the infected individuals. Following each round of multiplication, the infected cell is ruptured in a process known as egress to release a new generation of parasites. Egress is required for the disease to progress and is orchestrated by the parasite. The parasite sends out various molecules to puncture and destroy the membranes of the vacuole and the red blood cell. However, little is known about these molecules. In this work, we set out to identify these molecules by using genetic and proteomics approaches. We screened the molecules the parasite sends out during egress and identified a parasite enzyme called LCAT present in the vacuole. Our experiments found that mutant parasites that were unable to make LCAT clumped together and could not escape the infected cell properly. As a result, we saw a reduction in the rate at which these parasites spread through the red blood cells. Taken together, our findings suggest that P. falciparum needs LCAT to efficiently break out of red blood cells.

microbiology↗

bioRxiv2023

Comparative proteomics of vesicles essential for the egress of Plasmodium falciparum gametocytes from red blood cells

Transmission of malaria parasites to the mosquito is mediated by sexual precursor cells, the gametocytes. Upon entering the mosquito midgut, the gametocytes egress from the enveloping erythrocyte while passing through gametogenesis. Egress follows an inside-out mode during which the membrane of the parasitophorous vacuole ruptures prior to the erythrocyte membrane. Membrane rupture requires the exocytosis of specialized secretory vesicles of the parasites; i.e. the osmiophilic bodies (OBs) involved in rupturing the parasitophorous vacuole membrane, and vesicles (here termed g-exonemes) that harbour the perforin-like protein PPLP2 required for erythrocyte lysis. While several OB proteins are known, like G377 and MDV1/Peg3, the protein composition of the g-exonemes remains unclear. Here, we used high- resolution imaging and BioID methods to study the two types of egress vesicles in Plasmodium falciparum gametocytes. We show that OB exocytosis precedes discharge of the g-exonemes and that exocytosis of the g-exonemes, but not of the OBs, is calcium-sensitive. Further, the two types of vesicles exhibit distinct proteomes. In addition to known egress-related proteins, our analyses revealed novel components of OBs and g-exonemes, including proteins involved in vesicle trafficking. Our data provide insight into the immense molecular machinery required for the inside-out egress of P. falciparum gametocytes.

molecular biology↗

bioRxiv2022

A choline-releasing glycerophosphodiesterase essential for phosphatidylcholine biosynthesis and blood stage development in the malaria parasite

The malaria parasite Plasmodium falciparum synthesizes significant amounts of phospholipids to meet the demands of replication within red blood cells. De novo phosphatidylcholine (PC) biosynthesis via the Kennedy pathway is essential, requiring choline that is primarily sourced from host serum lysophosphatidylcholine (lysoPC). LysoPC also acts as an environmental sensor to regulate parasite sexual differentiation. Despite these critical roles for host lysoPC, the enzyme(s) involved in its breakdown to free choline for PC synthesis are unknown. Here we show that a parasite glycerophosphodiesterase (PfGDPD) is indispensable for blood stage parasite proliferation. Exogenous choline rescues growth of PfGDPD-null parasites, directly linking PfGDPD function to choline incorporation. Loss of PfGDPD reduces choline uptake from lysoPC, resulting in depletion of several PC species in the parasite, whilst purified PfGDPD releases choline from glycerophosphocholine in vitro. Our results identify PfGDPD as a choline-releasing glycerophosphodiesterase that mediates a critical step in PC biosynthesis and parasite survival.

microbiology↗

bioRxiv2021

Characterization of apicomplexan amino acid transporters (ApiATs) in the malaria parasite Plasmodium falciparum

During the symptomatic human blood phase, malaria parasites replicate within red blood cells. Parasite proliferation relies on the uptake of nutrients, such as amino acids, from the host cell and the blood plasma, requiring transport across multiple membranes. Amino acids are delivered to the parasite through the parasite surrounding vacuolar compartment by specialized nutrient-permeable channels of the erythrocyte membrane and the parasitophorous vacuole membrane (PVM). However, further transport of amino acid across the parasite plasma membrane (PPM) is currently not well characterized. In this study, we focused on a family of Apicomplexan amino acid transporters (ApiATs) that comprises five members in Plasmodium falciparum. First, we localized four of the PfApiATs at the PPM using endogenous GFP-tagging. Next, we applied reverse genetic approaches to probe into their essentiality during asexual replication and gametocytogenesis. Upon inducible knockdown and targeted gene disruption a reduced asexual parasite proliferation was detected for PfApiAT2 and PfApiAT4. Functional inactivation of individual PfApiATs targeted in this study had no effect on gametocyte development. Our data suggest that individual PfApiATs are partially redundant during asexual in vitro proliferation and fully redundant during gametocytogenesis of P. falciparum parasites. IMPORTANCEMalaria parasites live and multiply inside cells. To facilitate their extremely fast intracellular proliferation they hijack and transform their host cells. This also requires the active uptake of nutrients, such as amino acids, from the host cell and the surrounding environment through various membranes that are the consequence of the parasites intracellular lifestyle. In this manuscript we focus on a family of putative amino acid transporters termed ApiAT. We show expression and localization of four transporters in the parasite plasma membrane of Plasmodium falciparum-infected erythrocytes that represent one interface of the pathogen to its host cell. We probed into the impact of functional inactivation of individual transporters on parasite growth in asexual and sexual blood stages of P. falciparum and reveal that only two of them show a modest but significant reduction in parasite proliferation but no impact on gametocytogenesis pointing towards redundancy within this transporter family.

molecular biology↗

bioRxiv2021

Global analysis of putative phospholipases in the malaria parasite Plasmodium falciparum reveals critical factors for parasite proliferation

For its replication within red blood cells, the malaria parasite is highly dependent on correctly regulated lipid metabolism. Enzymes involved in lipid metabolic processes are therefore potential drug targets. We here provide a functional analysis of the 20 putative phospholipases that are expressed by asexual blood stages of Plasmodium falciparum. We reveal a high level of redundancy among members of this group, but using conditional mislocalization and gene disruption techniques we show that the phosphoinositide-specific phospholipase C (PF3D7_1013500) has a previously unrecognized essential role in intracellular parasite maturation. In addition, we demonstrate that the patatin-like phospholipase PF3D7_1358000 localizes to the mitochondrion. Parasites lacking this enzyme display a severe growth phenotype and defects in mitochondrial morphogenesis and function leading to hypersensitivity towards proguanil and inhibitors of the mitochondrial electron transport chain including atovaquone. This demonstrates that regulated mitochondrial lipid homeostasis is necessary for mitochondrial function and coordinated division during parasite multiplication.

microbiology↗

bioRxiv2021

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

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.

cell biology↗

bioRxiv2020

Structure-based identification and functional characterization of an essential lipocalin in the malaria parasite Plasmodium falciparum

Proteins of the lipocalin family are known to bind small hydrophobic ligands and are involved in various physiological processes ranging from lipid transport to oxidative stress responses. The genome of the malaria parasite Plasmodium falciparum contains a single protein PF3D7_0925900 with a lipocalin signature. Using crystallography and small-angle X-ray scattering, we show that the protein has a tetrameric structure of typical lipocalin monomers, hence we name it P. falciparum lipocalin (PfLCN), the first lipocalin structurally and functionally characterized in a single-celled eukaryote. We show that PfLCN is expressed in the intraerythrocytic stages of the parasite and localizes to the parasitophorous and food vacuoles. Conditional knockdown of PfLCN impairs parasite development, which can be rescued by treatment with the radical scavenger Trolox or by temporal inhibition of hemoglobin digestion. This suggests a key function of PfLCN in counteracting oxidative stress induced cell damage during multiplication of parasites within red blood cells.

cell biology↗