Inhibition of malaria and babesiosis parasites by putative red blood cell targeting small molecules.

Background: Chemotherapies for malaria and babesiosis frequently succumb to the emergence of pathogen-related drug-resistance. Host-targeted therapies are thought to be less susceptible to resistance but are seldom considered for treatment of these diseases.

Methods: Our overall objective was to systematically assess small molecules for host cell-targeting activity to restrict proliferation of intracellular parasites.

Comparative chemical genomics in species identifies the alkaline phosphatase PhoD as a determinant of antiparasitic resistance.

is an emerging zoonosis and widely distributed veterinary infection caused by 100+ species of parasites. The diversity of parasites and the lack of specific drugs necessitate the discovery of broadly effective antibabesials. Here, we describe a comparative chemogenomics (CCG) pipeline for the identification of conserved targets.

Acetoxy Group-Directed Regioselective C2 Alkenylation of Indoles via Pd-Ag Bimetallic Catalysis.

Regioselective C-H functionalizations of indoles reported to date with directing groups at C3 mainly rely on functional groups that are linked to the indole via C-C bonds. However, groups that are linked to the indole core by C-X linkages are also attractive due to the possibility of further modifications of the C-X bond. Herein, we report a 3-acetoxy directing group for the regioselective C2 alkenylation of indoles via a C-H activation-based, cross-dehydrogenative, oxidative Heck-type reaction.

Comparative chemical genomics in species identifies the alkaline phosphatase phoD as a novel determinant of resistance.

Unlabelled: is an emerging zoonosis and widely distributed veterinary infection caused by 100+ species of parasites. The diversity of parasites, coupled with the lack of potent inhibitors necessitates the discovery of novel conserved druggable targets for the generation of broadly effective antibabesials. Here, we describe a comparative chemogenomics (CCG) pipeline for the identification of novel and conserved targets.

Visible-light driven acetoxylation and dioxygenation of indoles electron donor-acceptor complexes.

A photoresponsive electron donor-acceptor (EDA) complex, formed by indole and hypervalent iodine such as diacetoxyiodobenzene (DAIB/PIDA, ), was detectable through absorption and emission spectroscopy. Irradiation of the EDA complex with visible light triggered photoinduced single electron transfer (SET) processes that were synthetically useful for the catalyst-free, regioselective acetylation of indoles. The photocatalytic reaction with excess DAIB was also used for the synthesis of isatins.

Babesia divergens egress from host cells is orchestrated by essential and druggable kinases and proteases.

Apicomplexan egress from host cells is fundamental to the spread of infection and is poorly characterized in spp., parasites of veterinary importance and emerging zoonoses. Through the use of video microscopy, transcriptomics and chemical genetics, we have implicated signaling, proteases and gliding motility as key drivers of egress by .

Organophotoredox-Catalyzed Cross-Dehydrogenative Sulfonamidation of Indoles and Other Heterocycles.

The visible light-triggered regioselective synthesis of 2-sulfonamidoindoles and other 2-sulfonamido heteroarenes is accomplished by the oxidative cross-dehydrogenative coupling of indoles (heteroarenes) with di--toluenesulfonamide or -aryl--toluenesulfonamides. The reaction was catalyzed by eosin-Y through a photoredox route. Detailed mechanistic studies based on control reactions, cyclic voltammetry, and fluorescence quenching have been reported for the elucidation of the mechanistic cycle and revealed that a nitrogen-centered radical is generated, followed by regioselective addition to the heteroarene.

Co-option of Plasmodium falciparum PP1 for egress from host erythrocytes.

Asexual proliferation of the Plasmodium parasites that cause malaria follows a developmental program that alternates non-canonical intraerythrocytic replication with dissemination to new host cells. We carried out a functional analysis of the Plasmodium falciparum homolog of Protein Phosphatase 1 (PfPP1), a universally conserved cell cycle factor in eukaryotes, to investigate regulation of parasite proliferation. PfPP1 is indeed required for efficient replication, but is absolutely essential for egress of parasites from host red blood cells.

Fussing About Fission: Defining Variety Among Mainstream and Exotic Apicomplexan Cell Division Modes.

Cellular reproduction defines life, yet our textbook-level understanding of cell division is limited to a small number of model organisms centered around humans. The horizon on cell division variants is expanded here by advancing insights on the fascinating cell division modes found in the Apicomplexa, a key group of protozoan parasites. The Apicomplexa display remarkable variation in offspring number, whether karyokinesis follows each S/M-phase or not, and whether daughter cells bud in the cytoplasm or bud from the cortex.

Targeting Plasmodium Proteases to Block Malaria Parasite Escape and Entry.

Proliferation of malaria parasites in a host requires mechanisms to spread between red blood cells (RBCs). We discuss here the implications for biology and antimalarial drug development of companion studies that establish the requirement of two Plasmodium spp. proteases of the plasmepsin family in parasite egress from, and invasion into, RBCs.

PfCDPK1 mediated signaling in erythrocytic stages of Plasmodium falciparum.

Calcium Dependent Protein Kinases are key effectors of calcium signaling in malaria parasite. PfCDPK1 is critical for asexual development of Plasmodium falciparum, but its precise function and substrates remain largely unknown. Using a conditional knockdown strategy, we here establish that this kinase is critical for the invasion of host erythrocytes.

Host Cell Tropism and Adaptation of Blood-Stage Malaria Parasites: Challenges for Malaria Elimination.

and account for most of the mortality and morbidity associated with malaria in humans. Research and control efforts have focused on infections caused by and , but have neglected other malaria parasite species that infect humans. Additionally, many related malaria parasite species infect nonhuman primates (NHPs), and have the potential for transmission to humans.

Plasmodium falciparum CRK4 directs continuous rounds of DNA replication during schizogony.

Plasmodium parasites, the causative agents of malaria, have evolved a unique cell division cycle in the clinically relevant asexual blood stage of infection. DNA replication commences approximately halfway through the intracellular development following invasion and parasite growth. The schizont stage is associated with multiple rounds of DNA replication and nuclear division without cytokinesis, resulting in a multinucleated cell.

Extensive Shared Chemosensitivity between Malaria and Babesiosis Blood-Stage Parasites.

The apicomplexan parasites that cause malaria and babesiosis invade and proliferate within erythrocytes. To assess the potential for common antiparasitic treatments, we measured the sensitivities of multiple species of Plasmodium and Babesia parasites to the chemically diverse collection of antimalarial compounds in the Malaria Box library. We observed that these parasites share sensitivities to a large fraction of the same inhibitors and we identified compounds with strong babesiacidal activity.

Using a Genetically Encoded Sensor to Identify Inhibitors of Toxoplasma gondii Ca2+ Signaling.

The life cycles of apicomplexan parasites progress in accordance with fluxes in cytosolic Ca(2+) Such fluxes are necessary for events like motility and egress from host cells. We used genetically encoded Ca(2+) indicators (GCaMPs) to develop a cell-based phenotypic screen for compounds that modulate Ca(2+) signaling in the model apicomplexan Toxoplasma gondii In doing so, we took advantage of the phosphodiesterase inhibitor zaprinast, which we show acts in part through cGMP-dependent protein kinase (protein kinase G; PKG) to raise levels of cytosolic Ca(2+) We define the pool of Ca(2+) regulated by PKG to be a neutral store distinct from the endoplasmic reticulum. Screening a library of 823 ATP mimetics, we identify both inhibitors and enhancers of Ca(2+) signaling.

Parasite Calcineurin Regulates Host Cell Recognition and Attachment by Apicomplexans.

Apicomplexans invade a variety of metazoan host cells through mechanisms involving host cell receptor engagement and secretion of parasite factors to facilitate cellular attachment. We find that the parasite homolog of calcineurin, a calcium-regulated phosphatase complex central to signal transduction in eukaryotes, also contributes to host cell invasion by the malaria parasite Plasmodium falciparum and related Toxoplasma gondii. Using reverse-genetic and chemical-genetic approaches, we determine that calcineurin critically regulates and stabilizes attachment of extracellular P.

Malaria. A forward genetic screen identifies erythrocyte CD55 as essential for Plasmodium falciparum invasion.

Efforts to identify host determinants for malaria have been hindered by the absence of a nucleus in erythrocytes, which precludes genetic manipulation in the cell in which the parasite replicates. We used cultured red blood cells derived from hematopoietic stem cells to carry out a forward genetic screen for Plasmodium falciparum host determinants. We found that CD55 is an essential host factor for P.

Host-parasite interactions that guide red blood cell invasion by malaria parasites.

Purpose Of Review: Malaria is caused by the infection and proliferation of parasites from the genus Plasmodium in red blood cells (RBCs). A free Plasmodium parasite, or merozoite, released from an infected RBC must invade another RBC host cell to sustain a blood-stage infection. Here, we review recent advances on RBC invasion by Plasmodium merozoites, focusing on specific molecular interactions between host and parasite.

Specific phosphorylation of the PfRh2b invasion ligand of Plasmodium falciparum.

Red blood cell invasion by the malaria parasite Plasmodium falciparum relies on a complex protein network that uses low and high affinity receptor-ligand interactions. Signal transduction through the action of specific kinases is a control mechanism for the orchestration of this process. In the present study we report on the phosphorylation of the CPD (cytoplasmic domain) of P.

Review of the mechanism of processive actin filament elongation by formins.

We review recent structural and biophysical studies of the mechanism of action of formins, proteins that direct the assembly of unbranched actin filaments for cytokinetic contractile rings and other cellular structures. Formins use free actin monomers to nucleate filaments and then remain bound to the barbed ends of these filaments as they elongate. In addition to variable regulatory domains, formins typically have formin homology 1 (FH1) and formin homology 2 (FH2) domains.

Energetic requirements for processive elongation of actin filaments by FH1FH2-formins.

Formin-homology (FH) 2 domains from formin proteins associate processively with the barbed ends of actin filaments through many rounds of actin subunit addition before dissociating completely. Interaction of the actin monomer-binding protein profilin with the FH1 domain speeds processive barbed end elongation by FH2 domains. In this study, we examined the energetic requirements for fast processive elongation.

A malaria parasite formin regulates actin polymerization and localizes to the parasite-erythrocyte moving junction during invasion.

Malaria parasites invade host cells using actin-based motility, a process requiring parasite actin filament nucleation and polymerization. Malaria and other apicomplexan parasites lack Arp2/3 complex, an actin nucleator widely conserved across eukaryotes, but do express formins, another type of actin nucleator. Here, we demonstrate that one of two malaria parasite formins, Plasmodium falciparum formin 1 (PfFormin 1), and its ortholog in the related parasite Toxoplasma gondii, follows the moving tight junction between the invading parasite and the host cell, which is the predicted site of the actomyosin motor that powers motility.

The role of the FH1 domain and profilin in formin-mediated actin-filament elongation and nucleation.

Background: Formin proteins nucleate actin filaments de novo and stay associated with the growing barbed end. Whereas the formin-homology (FH) 2 domains mediate processive association, the FH1 domains-in concert with the actin-monomer-binding protein profilin-increase the rate of barbed-end elongation. The mechanism by which this effect is achieved is not well understood.

Importance of a repetitive nine-residue sequence motif for intracellular stability and functional structure of a family I.3 lipase.

PML5 is a functional derivative of a family I.3 lipase from Pseudomonas sp. MIS38 and contains five repeats of a nine-residue sequence motif.