Biology subjects

Merino, E. F.

Publications and source records attributed to Merino, E. F..

4 recordsLinked to original sources

bioRxiv2021

Alkyne modified purines for assessing activation of Plasmodium vivax hypnozoites and growth of pre-erythrocytic and erythrocytic stages in Plasmodium spp.

Malaria is a major global health problem which predominantly afflicts developing countries. Although many antimalarial therapies are currently available, the protozoan parasite causing this disease, Plasmodium spp., continues to evade eradication efforts. One biological phenomenon hampering eradication efforts is the parasites ability to arrest development, transform into a drug-insensitive form, and then resume growth post-therapy. Currently, the mechanisms by which the parasite enters arrested development, or dormancy, and later recrudesces or reactivates to continue development, are unknown and the malaria field lacks techniques to study these elusive mechanisms. Since Plasmodium spp. salvage purines for DNA synthesis, we hypothesized that alkyne-containing purine nucleosides could be used to develop a DNA synthesis marker which could be used to investigate mechanisms behind dormancy. Using copper-catalyzed click chemistry methods, we observe incorporation of alkyne modified adenosine, inosine, and hypoxanthine in actively replicating asexual blood stages of P. falciparum and incorporation of modified adenosine in actively replicating liver stage schizonts of P. vivax. Notably, these modified purines were not incorporated in dormant liver stage hypnozoites, suggesting this marker could be used as a tool to differentiate replicating and non-replicating liver forms and, more broadly, a tool for advancing our understanding Plasmodium dormancy mechanisms.

microbiology↗

bioRxiv2019

Metabolic dependency of chorismate in Plasmodium falciparum

The shikimate pathway, a metabolic pathway absent in humans, is responsible for the production of chorismate, a branch point metabolite. In the malaria parasite, chorismate is postulated to be a direct precursor in the synthesis of p-aminobenzoic acid (folate biosynthesis), p-hydroxybenzoic acid (ubiquinone biosynthesis), menaquinone, and aromatic amino acids. While the potential value of the shikimate pathway as a drug target is debatable, the metabolic dependency of chorismate in P. falciparum remains unclear. Current evidence suggests that the main role of chorismate is folate biosynthesis despite ubiquinone biosynthesis being active and essential in the malaria parasite. Our goal in the present work was to expand our knowledge of the ubiquinone head group biosynthesis and its potential metabolic dependency on chorismate in P. falciparum. These data led us to further characterize the mechanism of action of MMV688345, a compound from the open-access \"Pathogen Box\" collection from Medicine for Malaria Venture. We systematically assessed the development of both asexual and sexual stages of P. falciparum in a defined medium in the absence of an exogenous supply of chorismate end-products and present biochemical evidence suggesting that the benzoquinone ring of ubiquinones in this parasite may be synthesized through a yet unidentified route.

biochemistry↗

bioRxiv2019

Metabolomic profiling reveals developmentally regulated biosynthesis of polyprenols and dolichols in the malaria parasite

The cis-polyisoprenoid lipids namely polyprenols, dolichols and their derivatives are linear polymers of several isoprene units. In eukaryotes, polyprenols and dolichols are synthesized as a mixture of four or more homologues of different length with one or two predominant species with sizes varying among organisms. Polyprenols have been hardly detectable in eukaryotic cells under normal conditions with the exception of plants and sporulating yeast. Our metabolomics studies revealed that cis-polyisoprenoids are more prevalent and diverse in the parasite Plasmodium falciparum than previously postulated as we uncovered active de novo biosynthesis and substantial levels of accumulation of polyprenols and dolichols of 15 to 19 isoprene units. A distinctive polyprenol and dolichol profile both within the intraerythrocytic asexual cycle and between asexual and gametocyte stages was also observed suggesting that cis-polyisoprenoid biosynthesis changes throughout parasites development. In addition, we confirmed the presence of an active cis-prenyltransferase (PfCPT) and that dolichol biosynthesis occurs via reduction of the polyprenol to dolichol by an active polyprenol reductase (PfPPRD) in the malaria parasite. Isotopic labeling and metabolomic analyses of a conditional mutant of PfCPT or PfPPRD suggest that polyprenols may be able to substitute dolichols in their biological functions when dolichol synthesis is impaired in Plasmodium.

biochemistry↗

bioRxiv2017

Suppressor screen connects HAD protein function to metabolic control in Plasmodium falciparum

In the malaria parasite Plasmodium falciparum, synthesis of isoprenoids from glycolytic intermediates is essential for survival. The antimalarial fosmidomycin (FSM) inhibits isoprenoid synthesis. In P. falciparum, we identify a loss-of-function mutation in HAD2 (PF3D7_1226300) as necessary for FSM resistance. Enzymatic characterization reveals that HAD2, a member of the haloacid dehalogenase-like hydrolase (HAD) superfamily, is a phosphatase. Harnessing a growth defect in resistant parasites, we select for suppression of HAD2-mediated FSM resistance and uncover hypomorphic suppressor mutations in the locus encoding the glycolytic enzyme phosphofructokinase (PFK9). Metabolic profiling demonstrates that FSM resistance is achieved via increased steady-state levels of MEP pathway and glycolytic intermediates and confirms reduced PFK9 function in the suppressed strains. We identify HAD2 as a novel regulator of malaria parasite metabolism and drug sensitivity and uncover PFK9 as a novel site of genetic metabolic plasticity in the parasite. Our study informs the biological functions of an evolutionarily conserved family of metabolic regulators and reveals a previously undescribed strategy by which malaria parasites adapt to cellular metabolic dysregulation.\n\nIMPORTANCEUnique and essential aspects of parasite metabolism are excellent targets for development of new antimalarials. An improved understanding of parasite metabolism and drug resistance mechanisms are urgently needed. The antibiotic fosmidomycin targets the synthesis of essential isoprenoid compounds from glucose and is a candidate for antimalarial development. Our study identifies a novel mechanism of drug resistance and further describes a family of metabolic regulators in the parasite. Using a novel forward genetic approach, we also uncover mutations that suppress drug resistance in the glycolytic enzyme PFK9. Thus, we identify an unexpected genetic mechanism of adaptation to metabolic insult that influences parasite fitness and tolerance to antimalarials.

microbiology↗