Biology subjects

Winzeler, E. A.

Publications and source records attributed to Winzeler, E. A..

5 recordsLinked to original sources

bioRxiv2018

Validation of the protein kinase PfCLK3 as a multi-stage cross species malarial drug target

The requirement for next generation anti-malarials to be both curative and transmission blockers necessitate the identification of molecular pathways essential for viability of both asexual and sexual parasite life stages. Here we identify a selective inhibitor to the Plasmodium falciparum protein kinase PfCLK3 which we use in combination with chemogenetics, whole genome sequencing and transcriptomics to validate PfCLK3 as a druggable target acting at multiple parasite life stages. Consistent with the proposed role of PfCLK3 as a regulator of RNA splicing, inhibition results in the down-regulation of >400 genes essential for parasite survival. Through this mechanism, blocking PfCLK3 activity not only results in rapid killing of asexual blood stage parasites but is also effective on sporozoites and gametocytes as well as showing parasiticidal activity in all Plasmodium species tested. Hence, our data establishes PfCLK3 as a target with the potential to deliver both symptomatic treatment and transmission blocking in malaria.

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↗

bioRxiv2017

Dual RNAseq shows the human mucosal immunity protein, MUC13, is a hallmark of Plasmodium exoerythrocytic infection

The exoerythrocytic stage of Plasmodium malaria infection is a critical window for prophylactic intervention. Using a genome-wide dual RNA sequencing of flow-sorted infected and uninfected hepatoma cells we identify the human mucosal immunity gene, Mucin13 (MUC13), as strongly upregulated during Plasmodium exoerythrocytic hepatic-stage infection. We confirm that MUC13 expression is upregulated in hepatoma cell lines and primary hepatocytes. In immunofluorescence assays, host MUC13 protein expression distinguishes infected cells from adjacent uninfected cells and shows similar colocalization with parasite biomarkers such as UIS4 and HSP70. We further show that localization patterns are species independent, distinguishing both P. berghei and P. vivax infected cells, and that MUC13 can be used to identify compounds that inhibit parasite replication in hepatocytes across all Human-infecting Plasmodium species. This data presents a novel interface of host-parasite interactions in Plasmodium, in that a component of host mucosal immunity is reprogrammed to assist the progression of infection.

microbiology↗

bioRxiv2017

Common Piezo1 allele in African populations causes xerocytosis and attenuates Plasmodium infection

Hereditary xerocytosis (HX) is thought to be a rare genetic condition characterized by red blood cell (RBC) dehydration with mild hemolysis. Gain-of-function (GOF) mutations in mechanosensitive Piezo1 ion channels are identified in HX patients. RBC dehydration is linked to reduced Plasmodium infection rates in vitro. We engineered a Piezo1 mouse model of HX and show that Plasmodium infection fails to cause experimental cerebral malaria in these mice. Furthermore, we identified a novel GOF human Piezo1 variant, E756del, present in a third of African population. Remarkably, RBCs from individuals carrying this allele are dehydrated and protected against Plasmodium infection in vitro. The presence of an HX-causing Piezo1 mutation at such high frequencies in African population is surprising, and suggests an association with malaria resistance.

physiology↗

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↗