408 research outputs found
Evidence-based annotation of the malaria parasite's genome using comparative expression profiling
No full textThis is the companion web site for publication:Zhou Y, Ramachandran V, Kumar KA, Westenberger S, Refour P, Zhou B, Li F, Young JA, Chen K, Plouffe D, Henson K, Nussenzweig V, Carlton J, Vinetz JM, Duraisingh MT, Winzeler EA. Evidence-based annotation of the malaria parasite's genome using comparative expression profiling. PLoS One. 2008 Feb 13;3(2):e1570. doi: 10.1371/journal.pone.0001570. PMID: 18270564; PMCID: PMC2215772.Download and unzip the file, open Py/index.html to browse the self-contained web site.</div
The Plasmodium falciparum sexual development transcriptome: a microarray analysis using ontology-based pattern identification
No full textThis is the companion web site for publication:Young JA, Fivelman QL, Blair PL, de la Vega P, Le Roch KG, Zhou Y, Carucci DJ, Baker DA, Winzeler EA. The Plasmodium falciparum sexual development transcriptome: a microarray analysis using ontology-based pattern identification. Mol Biochem Parasitol. 2005 Sep;143(1):67-79. doi: 10.1016/j.molbiopara.2005.05.007. PMID: 16005087.Download and unzip the file, open Gametocyte/index.html to browse the self-contained web site.</div
In silico discovery of transcription regulatory elements in Plasmodium falciparum
No full textThis is the companion web site for publication:Young JA, Johnson JR, Benner C, Yan SF, Chen K, Le Roch KG, Zhou Y, Winzeler EA. In silico discovery of transcription regulatory elements in Plasmodium falciparum. BMC Genomics. 2008 Feb 7;9:70. doi: 10.1186/1471-2164-9-70. PMID: 18257930; PMCID: PMC2268928.Download and unzip the file, open motif/index.html to browse the self-contained web site.</div
A systems-based analysis of Plasmodium vivax lifecycle transcription from human to mosquito
No full textThis is the companion web site for publication:Westenberger SJ, McClean CM, Chattopadhyay R, Dharia NV, Carlton JM, Barnwell JW, Collins WE, Hoffman SL, Zhou Y, Vinetz JM, Winzeler EA. A systems-based analysis of Plasmodium vivax lifecycle transcription from human to mosquito. PLoS Negl Trop Dis. 2010 Apr 6;4(4):e653. doi: 10.1371/journal.pntd.0000653. PMID: 20386602; PMCID: PMC2850316.Download and unzip the file, open Pv/index.html to browse the self-contained web site.</div
Discovery of Gene Function by Expression Profiling of the Malaria Parasite Life Cycle
No full textThis is the companion web site for publication:Le Roch KG, Zhou Y, Blair PL, Grainger M, Moch JK, Haynes JD, De La Vega P, Holder AA, Batalov S, Carucci DJ, Winzeler EA. Discovery of gene function by expression profiling of the malaria parasite life cycle. Science. 2003 Sep 12;301(5639):1503-8. doi: 10.1126/science.1087025. Epub 2003 Jul 31. PMID: 12893887.Download and unzip the file, open CellCycle/index.html to browse the self-contained web site.</div
A systematic map of genetic variation in Plasmodium falciparum
No full textThis is the companion web site for publication:Kidgell C, Volkman SK, Daily J, Borevitz JO, Plouffe D, Zhou Y, Johnson JR, Le Roch K, Sarr O, Ndir O, Mboup S, Batalov S, Wirth DF, Winzeler EA. A systematic map of genetic variation in Plasmodium falciparum. PLoS Pathog. 2006 Jun;2(6):e57. doi: 10.1371/journal.ppat.0020057. Epub 2006 Jun 23. Erratum in: PLoS Pathog. 2006 Aug;2(8):e96. PMID: 16789840; PMCID: PMC1480597.Download and unzip the file, open SFP/index.html to browse the self-contained web site.</div
Distinct physiological states of Plasmodium falciparum in malaria-infected patients
No full textThis is the companion web site for publication:Daily JP, Scanfeld D, Pochet N, Le Roch K, Plouffe D, Kamal M, Sarr O, Mboup S, Ndir O, Wypij D, Levasseur K, Thomas E, Tamayo P, Dong C, Zhou Y, Lander ES, Ndiaye D, Wirth D, Winzeler EA, Mesirov JP, Regev A. Distinct physiological states of Plasmodium falciparum in malaria-infected patients. Nature. 2007 Dec 13;450(7172):1091-5. doi: 10.1038/nature06311. Epub 2007 Nov 28. PMID: 18046333.Download and unzip the file, open Pv/index.html to browse the self-contained web site.</div
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Target Identification and Validation of the Imidazolopiperazine Class of Antimalarials
No full textThe antimalarial resistance arms race is alive and well. In 2022, the Plasmodium genus of parasites were responsible for 249 million cases of malaria, 608,000 of which were fatal (WHO, 2023). This increase in cases is attributed to lack of an effective vaccine and restricted access to preventative and treatment measures coming after the COVID-19 pandemic, but especially to the ongoing development of antimalarial resistant parasites. Thus, there is a dire need to create medicines that possess novel modes of action. And if malaria were ever to be eradicated, we need those that inhibit parasite growth at every stage of their life cycle. So, how do we know which compounds exhibit antimalarial activity in a novel way?This dissertation highlights and applies strategies of antimalarial target identification. Chapter 1 describes omics approaches: in-vitro evolution and whole genome analysis, proteomic affinity chromatography, cellular thermal shift assay, metabolomic profiling. Chapter 2 uses these methods on imidazolopiperazines, a new class of antimalarials that possess a novel mode of action and are active against multiple stages of the parasite life cycle. Previous work evolving parasites against the GNF179 analog only presented multidrug resistance mechanisms; metabolomic profiling with the KAF156 analog did not present clear perturbations. Using proteomic affinity chromatography, thermal shift assay, and surface plasmon resonance, we identify GNF179 interaction with a putative dynamin-like GTPase. Having only been predicted to be essential in parasites, we also confirm that it is an essential P. falciparum gene via conditional knockdown. Molecular docking and GTPase activity assay suggest imidazolopiperazine binding to the N-terminal GTPase domain, yet a nonsense mutation that removes residues from the C-terminal end confers resistance. In Chapter 3, we investigate the significance of the transmembrane domains and C-terminal tail of this GTPase protein to imidazolopiperazine interaction. Through this research, we can better guide imidazolopiperazine development and anticipate resistant alleles
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Know thy Enemy: Exploring Pathogenic Evolution, Resistance, and Virulence in Plasmodium falciparum and Leptospira interrogans
No full textWe are in a constant battle against pathogens. Millions of individuals die every year due to inadequate diagnosis and emerging resistance to current therapeutics. Improvements in diagnosis and treatment strategies are needed, but these can be hindered by a lack of knowledge regarding a particular pathogen’s molecular and resistance mechanisms. Plasmodium and Leptospira are two such examples of widespread pathogenic organisms whose genomes are still poorly understood and additional knowledge is desperately needed to improve therapeutic strategies. This dissertation presents two evolution-based strategies that when coupled with whole genome sequencing can be used to identify virulence and resistance associated genes. These include a “forward” approach, which studies the development of resistance, and a “reverse” approach, which examines the loss of virulence. In Chapter 2, the forward approach is used to examine novel targetable pathways in Plasmodium falciparum. Selectively evolving resistance to 50 novel antimalarial compounds, we successfully identify potential targets to 21 compounds and eight novel gene targets. Additionally, this chapter examines resistance development patterns against the compound set, and identifies fast-killing compounds may result in a slower onset of clinical resistance. Chapter 3 focuses on PfCARL, one potential target identified in Chapter 2, which has been previously described as the target for KAF156, a drug currently in clinical trials. Our data demonstrate that pfcarl mutations confer resistance to two distinct compound classes – benzimidazolyl piperidines and imidazolopiperazines. However, these two classes appear to have different timing of action in the asexual blood stage and different potencies against the liver and sexual blood stages, suggesting pfcarl is a multidrug resistance gene rather than a common target. Finally, using the reverse approach, Chapter 4 identifies virulence-related genes in Leptospira by observing cumulative genomic changes occurring after serial in vitro passaging of a highly virulent Leptospira interrogans strain into a nearly avirulent isogenic derivative. Comparison between these two polyclonal strains identifies 15 non-synonymous single nucleotide variant (nsSNV) alleles that increased in frequency and 19 that decreased. These frequency changes likely contribute to the loss of virulence, and suggest new virulence-associated genes whose role in Leptospira pathogenesis should be further studied
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Drug Target Discovery Using Designer Drug Sensitive Yeast
No full textDetermining the protein target(s) and mechanism(s) of drug candidates found in phenotypic screens is critical to subsequent structure-activity-based development and optimization, but existing methods for target identification are limited. Here we present a method that applies directed evolution to a genetically engineered, drug sensitive Saccharomyces cerevisiae strain. Whole genome sequencing of yeast clones that have evolved drug resistance, in concert with in vitro cell free assays and computer modeling, can be a useful tool for target identification and binding site characterization.To demonstrate the ease and utility of this method, we applied it to the identification of the molecular target and binding site of a range of cytotoxic molecular compounds with activity against eukaryotic pathogens and human cancers. These studies include known drug target combinations, as well as application to experimental compounds with unknown drug targets. As proof of concept, the method correctly identified the precise binding pocket of the protein synthesis inhibitor, cycloheximide, as the ribosomal protein Rpl28. We also correctly identified topoisomerase II inhibitor as the target of the human cancer chemotherapeutic, etoposide.We next used the method to identify novel drug target combinations, which were then validated using a combination of genetic, biochemical, structural and chemical structure activity relationships (SAR)-based assays. We identified a p-type ATPase, ScPma1, as the target of the spiroindolone antimalarials, of which KAE609 is currently in stage 2b clinical trials. We determined that the pre- clinical phenyl-amino-methyl-quinolinols (PAMQ) antimalarials inhibit the cyclic AMP signaling pathway, a mechanism of action that is different from existing commercial antimicrobials. We also demonstrated that MMV001239, a compound with antitrypanosomal activity, targets ScErg11, the yeast homolog of the T, cruzi Cyp51p, and a protein crucial for ergosterol biosynthesis. Taken together, our approach expands on the number of tools available for analyzing compound-target interactions and can be applied to studies of other eukaryotic antimicrobials and chemotherapeutics
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