An experimental target-based platform in yeast for screening Plasmodium vivax deoxyhypusine synthase inhibitors.

The enzyme deoxyhypusine synthase (DHS) catalyzes the first step in the post-translational modification of the eukaryotic translation factor 5A (eIF5A). This is the only protein known to contain the amino acid hypusine, which results from this modification. Both eIF5A and DHS are essential for cell viability in eukaryotes, and inhibiting DHS is a promising strategy to develop new therapeutic alternatives.

Identification of potential inhibitors of casein kinase 2 alpha of with potent activity.

Drug resistance to commercially available antimalarials is a major obstacle in malaria control and elimination, creating the need to find new antiparasitic compounds with novel mechanisms of action. The success of kinase inhibitors for oncological treatments has paved the way for the exploitation of protein kinases as drug targets in various diseases, including malaria. Casein kinases are ubiquitous serine/threonine kinases involved in a wide range of cellular processes such as mitotic checkpoint signaling, DNA damage response, and circadian rhythm.

Yeast Double Transporter Gene Deletion Library for Identification of Xenobiotic Carriers in Low or High Throughput.

The routes of uptake and efflux should be considered when developing new drugs so that they can effectively address their intracellular targets. As a general rule, drugs appear to enter cells via protein carriers that normally carry nutrients or metabolites. A previously developed pipeline that searched for drug transporters using Saccharomyces cerevisiae mutants carrying single-gene deletions identified import routes for most compounds tested.

Violacein-Induced Chaperone System Collapse Underlies Multistage Antiplasmodial Activity.

Antimalarial drugs with novel modes of action and wide therapeutic potential are needed to pave the way for malaria eradication. Violacein is a natural compound known for its biological activity against cancer cells and several pathogens, including the malaria parasite, (Pf). Herein, using chemical genomic profiling (CGP), we found that violacein affects protein homeostasis.

Structural features and development of an assay platform of the parasite target deoxyhypusine synthase of Brugia malayi and Leishmania major.

Deoxyhypusine synthase (DHS) catalyzes the first step of the post-translational modification of eukaryotic translation factor 5A (eIF5A), which is the only known protein containing the amino acid hypusine. Both proteins are essential for eukaryotic cell viability, and DHS has been suggested as a good candidate target for small molecule-based therapies against eukaryotic pathogens. In this work, we focused on the DHS enzymes from Brugia malayi and Leishmania major, the causative agents of lymphatic filariasis and cutaneous leishmaniasis, respectively.

Chemical Genomic Profiling Unveils the in Vitro and in Vivo Antiplasmodial Mechanism of Açaí ( Mart.) Polyphenols.

Malaria remains a major detrimental parasitic disease in the developing world, with more than 200 million cases annually. Widespread drug-resistant parasite strains push for the development of novel antimalarial drugs. Plant-derived natural products are key sources of antimalarial molecules.

Antiplasmodial and trypanocidal activity of violacein and deoxyviolacein produced from synthetic operons.

Background: Violacein is a deep violet compound that is produced by a number of bacterial species. It is synthesized from tryptophan by a pathway that involves the sequential action of 5 different enzymes (encoded by genes vioA to vioE). Violacein has antibacterial, antiparasitic, and antiviral activities, and also has the potential of inducing apoptosis in certain cancer cells.

Plasmodium dihydrofolate reductase is a second enzyme target for the antimalarial action of triclosan.

Malaria, caused by parasites of the genus Plasmodium, leads to over half a million deaths per year, 90% of which are caused by Plasmodium falciparum. P. vivax usually causes milder forms of malaria; however, P.

Yeast-Based High-Throughput Screens to Identify Novel Compounds Active against Brugia malayi.

Background: Lymphatic filariasis is caused by the parasitic worms Wuchereria bancrofti, Brugia malayi or B. timori, which are transmitted via the bites from infected mosquitoes. Once in the human body, the parasites develop into adult worms in the lymphatic vessels, causing severe damage and swelling of the affected tissues.

Cheaper faster drug development validated by the repositioning of drugs against neglected tropical diseases.

There is an urgent need to make drug discovery cheaper and faster. This will enable the development of treatments for diseases currently neglected for economic reasons, such as tropical and orphan diseases, and generally increase the supply of new drugs. Here, we report the Robot Scientist 'Eve' designed to make drug discovery more economical.

Identification and deconvolution of cross-resistance signals from antimalarial compounds using multidrug-resistant Plasmodium falciparum strains.

Plasmodium falciparum, the most deadly agent of malaria, displays a wide variety of resistance mechanisms in the field. The ability of antimalarial compounds in development to overcome these must therefore be carefully evaluated to ensure uncompromised activity against real-life parasites. We report here on the selection and phenotypic as well as genotypic characterization of a panel of sensitive and multidrug-resistant P.

Yeast-based automated high-throughput screens to identify anti-parasitic lead compounds.

We have developed a robust, fully automated anti-parasitic drug-screening method that selects compounds specifically targeting parasite enzymes and not their host counterparts, thus allowing the early elimination of compounds with potential side effects. Our yeast system permits multiple parasite targets to be assayed in parallel owing to the strains' expression of different fluorescent proteins. A strain expressing the human target is included in the multiplexed screen to exclude compounds that do not discriminate between host and parasite enzymes.

The promiscuous binding of pharmaceutical drugs and their transporter-mediated uptake into cells: what we (need to) know and how we can do so.

A recent paper in this journal sought to counter evidence for the role of transport proteins in effecting drug uptake into cells, and questions that transporters can recognize drug molecules in addition to their endogenous substrates. However, there is abundant evidence that both drugs and proteins are highly promiscuous. Most proteins bind to many drugs and most drugs bind to multiple proteins (on average more than six), including transporters (mutations in these can determine resistance); most drugs are known to recognise at least one transporter.

Genome-wide assessment of the carriers involved in the cellular uptake of drugs: a model system in yeast.

Background: The uptake of drugs into cells has traditionally been considered to be predominantly via passive diffusion through the bilayer portion of the cell membrane. The recent recognition that drug uptake is mostly carrier-mediated raises the question of which drugs use which carriers.

Results: To answer this, we have constructed a chemical genomics platform built upon the yeast gene deletion collection, using competition experiments in batch fermenters and robotic automation of cytotoxicity screens, including protection by 'natural' substrates.

Functional expression of parasite drug targets and their human orthologs in yeast.

Background: The exacting nutritional requirements and complicated life cycles of parasites mean that they are not always amenable to high-throughput drug screening using automated procedures. Therefore, we have engineered the yeast Saccharomyces cerevisiae to act as a surrogate for expressing anti-parasitic targets from a range of biomedically important pathogens, to facilitate the rapid identification of new therapeutic agents.

Methodology/principal Findings: Using pyrimethamine/dihydrofolate reductase (DHFR) as a model parasite drug/drug target system, we explore the potential of engineered yeast strains (expressing DHFR enzymes from Plasmodium falciparum, P.

Contributions of Saccharomyces cerevisiae to understanding mammalian gene function and therapy.

Due to its genetic tractability and ease of manipulation, the yeast Saccharomyces cerevisiae has been extensively used as a model organism to understand how eukaryotic cells grow, divide, and respond to environmental changes. In this chapter, we reasoned that functional annotation of novel genes revealed by sequencing should adopt an integrative approach including both bioinformatics and experimental analysis to reveal functional conservation and divergence of complexes and pathways. The techniques and resources generated for systems biology studies in yeast have found a wide range of applications.

Identification of putative regulatory upstream ORFs in the yeast genome using heuristics and evolutionary conservation.

Background: The translational efficiency of an mRNA can be modulated by upstream open reading frames (uORFs) present in certain genes. A uORF can attenuate translation of the main ORF by interfering with translational reinitiation at the main start codon. uORFs also occur by chance in the genome, in which case they do not have a regulatory role.

The Bre5/Ubp3 ubiquitin protease complex from budding yeast contributes to the cellular response to DNA damage.

The ubiquitination status of proteins can control numerous aspects of protein function through targeted destruction or by altering protein-protein interactions, subcellular localization, or enzymatic activity. In addition to enzymes that mediate the conjugation of ubiquitin moieties to target proteins, there are enzymes that catalyze the removal of ubiquitin, termed ubiquitin proteases. One such ubiquitin protease, Ubp3, exists in a complex with a partner protein: Bre5.

Tails of histones in DNA double-strand break repair.

DNA double-strand breaks (DSBs) are, arguably, the most deleterious form of DNA damage. An increasing body of evidence points to the inaccurate or inefficient repair of DSBs as a key step in tumorigenesis. Therefore, it is of great importance to understand the processes by which DSBs are detected and repaired.

Rck1 and Rck2 MAPKAP kinases and the HOG pathway are required for oxidative stress resistance.

We demonstrate a role in oxidative and metal stress resistance for the MAPK-activated protein kinases Rck1 and Rck2 in Saccharomyces cerevisiae. We show that Hog1 is robustly phosphorylated in a Pbs2-dependent way during oxidative stress, and that Rck2 also is phosphorylated under these circumstances. Hog1 concentrates in the nucleus in oxidative stress.