Biology subjects

Goldberg, D. E.

Publications and source records attributed to Goldberg, D. E..

5 recordsLinked to original sources

bioRxiv2018

Diverse chemical compounds target Plasmodium falciparum plasma membrane lipid homeostasis

Lipid homeostasis is essential for the maintenance of life. We previously reported that disruptions of the parasite Na+ homeostasis via inhibition of PfATP4 resulted in elevated cholesterol within the parasite plasma membrane as assessed by saponin sensitivity. A large number of compounds have been shown to target the parasite Na+ homeostasis. We therefore screened the same collection of 800 compounds to identify chemotypes that disrupted the parasite plasma membrane lipid homeostasis. Here, we show that the compounds disrupting parasite Na+ homeostasis also induced saponin sensitivity, an indication of parasite lipid homeostasis disruption. Remarkably, 13 compounds were identified that altered plasma membrane lipid composition independent of Na+ homeostasis disruption. Further studies suggest that these compounds target the Plasmodium falciparum Niemann-Pick Type C1-Related (PfNCR1) protein, which is hypothesized to be involved in maintaining plasma membrane lipid composition. PfNCR1, like PfATP4, appears to be targeted by multiple chemotypes with potential for drug discovery.

microbiology↗

bioRxiv2018

Assessment of biological role and insight into druggability of the Plasmodium falciparum protease plasmepsin V

Upon infection of a red blood cell (RBC), the malaria parasite Plasmodium falciparum drastically remodels its host by exporting hundreds of proteins into the RBC cytosol. This program of protein export is essential for parasite survival, hence there is interest in export-related proteins as potential drug targets. One proposed target is plasmepsin V (PMV), an aspartic protease that cleaves export-destined proteins in the parasite ER at a motif called the Plasmodium export element (PEXEL). This cleavage is essential for effector export across the vacuolar membrane. Despite long-standing interest in PMV, functional studies have been hindered by the failure of current technologies to produce a regulatable lethal depletion of PMV. To overcome this technical barrier, we designed a facile system for stringent post-transcriptional regulation, allowing a tightly controlled, tunable knockdown of PMV. Under maximal knockdown conditions, parasite growth was arrested, validating PMV as essential for parasite survival in RBCs. We found that PMV levels had to be dramatically depleted to affect parasite growth, suggesting that the parasite maintains this enzyme in substantial excess. This has important implications for antimalarial development. Additionally, we found that PMV-depleted parasites arrest immediately after invasion of the host cell, suggesting that PMV has an unappreciated role in early development that is distinct from its previously reported role in protein export in later-stage parasites.\n\nImportanceMalaria is endemic to large swaths of the developing world, causing nearly 500,000 deaths each year. While infection can be treated with antimalarial drugs, resistance continues to emerge to frontline antimalarials, spurring calls for new drugs and targets to feed the drug development pipeline. One proposed target is the aspartic protease plasmepsin V (PMV) that processes exported proteins, enabling the export program that remodels the host cell. This work uses facile genetic tools to produce lethal depletion of PMV, validating it as a drug target and showing that PMV is made in substantial excess in blood-stage parasites. Unexpectedly, PMV depletion leads to parasite death immediately after invasion of RBCs, distinct from other disruptions of the export pathway. This suggests that PMV inhibitors could lead to relatively rapid parasite death, and that PMV has additional unexplored role(s) during RBC infection.

microbiology↗

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↗

bioRxiv2018

Evaluation of current and emerging anti-malarial medicines for inhibition of Toxoplasma gondii growth in vitro

Toxoplasma gondii is a common zoonotic infection of humans and estimates indicate that 1-2 billion people are chronically infected. Although largely asymptomatic, chronic infection poses risk of serious disease due to reactivation should immunity decline. Current therapies for toxoplasmosis only control acute infection caused by actively proliferating tachyzoites but do not eradicate the chronic tissue cyst stages. As well, there are considerable adverse side effects of the most commonly used therapy of combined sulfadiazine and pyrimethamine. Targeting the folate pathway is also an effective treatment for malaria, caused by the related parasites Plasmodium spp., suggesting common agents might be used to treat both infections. Here we evaluated currently approved and newly emerging medicines for malaria to determine if such compounds might also prove useful for treating toxoplasmosis. Surprisingly, the majority of anti-malarial compounds being used currently or in development for treatment of malaria were only modestly effective at inhibiting in vitro growth of T. gondii tachyzoites. These findings suggest that many essential processes in P. falciparum that are targeted by anti-malarial compounds are either divergent, or non-essential in T. gondii, thus limiting options for repurposing of current antimalarial medicines for toxoplasmosis.

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↗