Biology subjects

Istvan, E. S.

Publications and source records attributed to Istvan, E. S..

4 recordsLinked to original sources

bioRxiv2026

Plasmodium falciparum Niemann-Pick Type C1-Related protein relies on its physicochemical properties for membrane contact site localization required for cholesterol homeostasis

The Niemann-Pick Type C1-Related protein of the malaria parasite Plasmodium falciparum, PfNCR1, is a promising anti-malarial drug target facilitating cholesterol homeostasis at the interface of the malaria parasite with its host-red blood cell. PfNCR1 is localized to otherwise functionally uncharacterized regions covering [~]half of the host-parasite interface. These regions are defined by exceptionally narrow membrane contact sites, leaving only [~]3-4 nm vertical aqueous space in between the membranes. Determining the origin and functional consequence of localization to the closely apposed membrane is central for our understanding of PfNCR1 as drug target but also offers a window into the mechanism of the group of homologous proteins, associated with congenital conditions and cancer. Here we define the mechanism of PfNCR1s membrane contact site localization and its implication for cholesterol transport. We identified a 141 amino acid long (amphipathic) helix - linker - (amphipathic) helix domain ("HLH domain") unique to Plasmodium spp., that is necessary for efficient localization of PfNCR1 to the narrow membrane contact sites. Mechanistically, we show that this localization relies on the HLH domains physicochemical properties. GPI-anchoring the isolated HLH domain or a version of the HLH domain in which the helices are replaced by the amphipathic helix of human ATG3 are sufficient to target a fluorescent protein to regions of endogenous PfNCR1. Functionally, we demonstrate that the degree of PfNCR1 localization to narrow contact sites qualitatively correlates with its ability to maintain cholesterol homeostasis, linking PfNCR1s membrane contact to its recognized transport function. Collectively, the results establish the HLH domain as key element for PfNCR1s localization and effectiveness in cholesterol transport while also opening avenues to probe the narrow membrane contact site regions with engineered proteins.

microbiology↗

bioRxiv2024

The Plasmodium falciparum NCR1 membrane protein is a novel antimalarial target that exports cholesterol to maintain membrane homeostasis

Malaria is an extremely devastating parasitic infection that kills over half a million people each year. It is the leading cause of death in many developing countries, in part, due to a lack of resources and readily available therapeutics. Unfortunately, the most prevalent and deadliest causative agent of malaria, Plasmodium falciparum, has developed resistance to nearly all currently available antimalarial drugs. The P. falciparum Niemann-Pick Type C1-related (PfNCR1) transporter has been identified as a druggable target, as it is required for membrane homeostasis of the parasite. However, the structure and detailed molecular mechanism of this membrane protein are not yet available. Here we present three structures of PfNCR1 both in the absence and presence of the functional inhibitor MMV009108 at resolutions between 2.98 [A] and 3.81 [A] using single-particle cryo-electron microscopy (cryo-EM). The data suggest that PfNCR1 binds cholesterol and forms a cholesterol transport tunnel to modulate the composition of the parasite plasma membrane. Cholesterol efflux assays substantiate this as they show that PfNCR1 is an exporter capable of extruding cholesterol from the membrane. Additionally, the inhibition mechanism of MMV009108 appears to be due to a direct blockage of PfNCR1, preventing this transporter from shuttling cholesterol.

biochemistry↗

bioRxiv2018

Plasmodium falciparum Niemann-Pick Type C1-Related Protein is a Druggable Target Required for Parasite Membrane Homeostasis

Plasmodium parasites possess a protein with homology to Niemann-Pick Type C1 proteins (Plasmodium falciparum Niemann-Pick Type C1-Related protein, PfNCR1). We isolated parasites with resistance-conferring mutations in PfNCR1 during selections with three diverse small-molecule antimalarial compounds and show that the mutations are causative for compound resistance. PfNCR1 protein knockdown results in severely attenuated growth and confers hypersensitivity to the compounds. Compound treatment or protein knockdown leads to increased sensitivity of the parasite plasma membrane (PPM) to the amphipathic glycoside saponin and engenders digestive vacuoles (DVs) that are small and malformed. Immuno-electron microscopy and split-GFP experiments localize PfNCR1 to the PPM. Our experiments show that PfNCR1 activity is critically important for the composition of the PPM and is required for DV biogenesis, suggesting PfNCR1 as a novel antimalarial drug target.

microbiology↗

bioRxiv2017

Mapping The Malaria Parasite Drug-Able Genome Using In Vitro Evolution And Chemogenomics

Chemogenetic characterization through in vitro evolution combined with whole genome analysis is a powerful tool to discover novel antimalarial drug targets and identify drug resistance genes. Our comprehensive genome analysis of 262 Plasmodium falciparum parasites treated with 37 diverse compounds reveals how the parasite evolves to evade the action of small molecule growth inhibitors. This detailed data set revealed 159 gene amplifications and 148 nonsynonymous changes in 83 genes which developed during resistance acquisition. Using a new algorithm, we show that gene amplifications contribute to 1/3 of drug resistance acquisition events. In addition to confirming known multidrug resistance mechanisms, we discovered novel multidrug resistance genes. Furthermore, we identified promising new drug target-inhibitor pairs to advance the malaria elimination campaign, including: thymidylate synthase and a benzoquinazolinone, farnesyltransferase and a pyrimidinedione, and a dipeptidylpeptidase and an arylurea. This deep exploration of the P. falciparum resistome and drug-able genome will guide future drug discovery and structural biology efforts, while also advancing our understanding of resistance mechanisms of the deadliest malaria parasite.\n\nOne Sentence SummaryWhole genome sequencing reveals how Plasmodium falciparum evolves resistance to diverse compounds and identifies new antimalarial drug targets.

genomics↗