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International Journal of Molecular... Jan 2022Malaria parasites require multiple phosphorylation and dephosphorylation steps to drive signaling pathways for proper differentiation and transformation. Several protein...
Malaria parasites require multiple phosphorylation and dephosphorylation steps to drive signaling pathways for proper differentiation and transformation. Several protein phosphatases, including protein phosphatase 1 (PP1), one of the main dephosphorylation enzymes, have been shown to be indispensable for the life cycle. The catalytic subunit of PP1 (PP1c) participates in cellular processes via dynamic interactions with a vast number of binding partners that contribute to its diversity of action. In this study, we used transgenic parasite strains stably expressing PP1c or its inhibitor 2 (I2) tagged with mCherry, combined with the mCherry affinity pulldown of proteins from asexual and sexual stages, followed by mass spectrometry analyses. Mapped proteins were used to identify interactomes and to cluster functionally related proteins. Our findings confirm previously known physical interactions of PP1c and reveal enrichment of common biological processes linked to cellular component assembly in both schizonts and gametocytes to biosynthetic processes/translation in schizonts and to protein transport exclusively in gametocytes. Further, our analysis of PP1c and I2 interactomes revealed that nuclear export mediator factor and peptidyl-prolyl cis-trans isomerase, suggested to be essential in , could be potential targets of the complex PP1c/I2 in both asexual and sexual stages. Our study emphasizes the adaptability of PP1 and provides a fundamental study of the protein interaction landscapes involved in a myriad of events in , suggesting why it is crucial to the parasite and a source for alternative therapeutic strategies.
Topics: Animals; Binding Sites; Chromatography, Liquid; Life Cycle Stages; Malaria; Male; Mice; Organisms, Genetically Modified; Plasmodium berghei; Protein Domains; Protein Interaction Maps; Protein Phosphatase 1; Proteins; Proteomics; Protozoan Proteins; Tandem Mass Spectrometry
PubMed: 35162991
DOI: 10.3390/ijms23031069 -
Scientific Reports Jan 2021The present study aimed to evaluate the effects of dexamethasone on the redox status, parasitemia evolution, and survival rate of Plasmodium berghei-infected mice....
The present study aimed to evaluate the effects of dexamethasone on the redox status, parasitemia evolution, and survival rate of Plasmodium berghei-infected mice. Two-hundred and twenty-five mice were infected with Plasmodium berghei and subjected to stimulation or inhibition of NO synthesis. The stimulation of NO synthesis was performed through the administration of L-arginine, while its inhibition was made by the administration of dexamethasone. Inducible NO synthase (iNOS) inhibition by dexamethasone promoted an increase in the survival rate of P. berghei-infected mice, and the data suggested the participation of oxidative stress in the brain as a result of plasmodial infection, as well as the inhibition of brain NO synthesis, which promoted the survival rate of almost 90% of the animals until the 15th day of infection, with possible direct interference of ischemia and reperfusion syndrome, as seen by increased levels of uric acid. Inhibition of brain iNOS by dexamethasone caused a decrease in parasitemia and increased the survival rate of infected animals, suggesting that NO synthesis may stimulate a series of compensatory redox effects that, if overstimulated, may be responsible for the onset of severe forms of malaria.
Topics: Animals; Arginine; Dexamethasone; Enzyme Inhibitors; Malaria; Male; Mice; Nitric Oxide; Parasitemia; Plasmodium berghei
PubMed: 33514836
DOI: 10.1038/s41598-021-82032-7 -
Proceedings of the National Academy of... Mar 2021Despite the critical role of sporozoites in malaria transmission, we still know little about the mechanisms underlying their development in mosquitoes. Here, we use...
Despite the critical role of sporozoites in malaria transmission, we still know little about the mechanisms underlying their development in mosquitoes. Here, we use single-cell RNA sequencing to characterize the gene expression profiles of 16,038 sporozoites isolated throughout their development from midgut oocysts to salivary glands, and from forced salivation experiments. Our results reveal a succession of tightly regulated changes in gene expression occurring during the maturation of sporozoites and highlight candidate genes that could play important roles in oocyst egress, sporozoite motility, and the mechanisms underlying the invasion of mosquito salivary glands and mammalian hepatocytes. In addition, the single-cell data reveal extensive transcriptional heterogeneity among parasites isolated from the same anatomical site, suggesting that development in mosquitoes is asynchronous and regulated by intrinsic as well as environmental factors. Finally, our analyses show a decrease in transcriptional activity preceding the translational repression observed in mature sporozoites and associated with their quiescent state in salivary glands, followed by a rapid reactivation of the transcriptional machinery immediately upon salivation.
Topics: Animals; Anopheles; Gene Expression Regulation; Mice; Plasmodium berghei; Salivary Glands; Sporozoites; Transcription, Genetic
PubMed: 33653959
DOI: 10.1073/pnas.2023438118 -
Parasites & Vectors Jan 2017Plasmodium ookinete surface proteins as post-fertilization target antigens are potential malaria transmission-blocking vaccine (TBV) candidates. Putative secreted...
BACKGROUND
Plasmodium ookinete surface proteins as post-fertilization target antigens are potential malaria transmission-blocking vaccine (TBV) candidates. Putative secreted ookinete protein 25 (PSOP25) is a highly conserved ookinete surface protein, and has been shown to be a promising novel TBV target. Here, we further investigated the TBV activities of the full-length recombinant PSOP25 (rPSOP25) protein in Plasmodium berghei, and characterized the potential functions of PSOP25 during the P. berghei life-cycle.
METHODS
We expressed the full-length P. berghei PSOP25 protein in a prokaryotic expression system, and developed polyclonal mouse antisera and a monoclonal antibody (mAb) against the recombinant protein. Indirect immunofluorescence assay (IFA) and Western blot were used to test the specificity of antibodies. The transmission-blocking (TB) activities of antibodies were evaluated by the in vitro ookinete conversion assay and by direct mosquito feeding assay (DFA). Finally, the function of PSOP25 during Plasmodium development was studied by deleting the psop25 gene.
RESULTS
Both polyclonal mouse antisera and anti-rPSOP25 mAb recognized the PSOP25 proteins in the parasites, and IFA showed the preferential expression of PSOP25 on the surface of zygotes, retorts and mature ookinetes. In vitro, these antibodies significantly inhibited ookinetes formation in an antibody concentration-dependent manner. In DFA, mice immunized with the rPSOP25 and those receiving passive transfer of the anti-rPSOP25 mAb reduced the prevalence of mosquito infection by 31.2 and 26.1%, and oocyst density by 66.3 and 63.3%, respectively. Genetic knockout of the psop25 gene did not have a detectable impact on the asexual growth of P. berghei, but significantly affected the maturation of ookinetes and the formation of midgut oocysts.
CONCLUSIONS
The full-length rPSOP25 could elicit strong antibody response in mice. Polyclonal and monoclonal antibodies against PSOP25 could effectively block the formation of ookinetes in vitro and transmission of the parasites to mosquitoes. Genetic manipulation study indicated that PSOP25 is required for ookinete maturation in P. berghei. These results support further testing of the PSOP25 orthologs in human malaria parasites as promising TBV candidates.
Topics: Animals; Antibodies, Protozoan; Antigens, Protozoan; Disease Models, Animal; Disease Transmission, Infectious; Gene Deletion; Immunization, Passive; Malaria; Malaria Vaccines; Mice; Plasmodium berghei; Protozoan Proteins
PubMed: 28057055
DOI: 10.1186/s13071-016-1932-4 -
Nature Communications Feb 2024A novel cellular response of midgut progenitors (stem cells and enteroblasts) to Plasmodium berghei infection was investigated in Anopheles stephensi. The presence of...
A novel cellular response of midgut progenitors (stem cells and enteroblasts) to Plasmodium berghei infection was investigated in Anopheles stephensi. The presence of developing oocysts triggers proliferation of midgut progenitors that is modulated by the Jak/STAT pathway and is proportional to the number of oocysts on individual midguts. The percentage of parasites in direct contact with enteroblasts increases over time, as progenitors proliferate. Silencing components of key signaling pathways through RNA interference (RNAi) that enhance proliferation of progenitor cells significantly decreased oocyst numbers, while limiting proliferation of progenitors increased oocyst survival. Live imaging revealed that enteroblasts interact directly with oocysts and eliminate them. Midgut progenitors sense the presence of Plasmodium oocysts and mount a cellular defense response that involves extensive proliferation and tissue remodeling, followed by oocysts lysis and phagocytosis of parasite remnants by enteroblasts.
Topics: Animals; Parasites; Janus Kinases; STAT Transcription Factors; Signal Transduction; Plasmodium; Malaria; Anopheles; Oocysts; Stem Cells; Plasmodium berghei
PubMed: 38365823
DOI: 10.1038/s41467-024-45550-2 -
Composition and organization of kinetochores show plasticity in apicomplexan chromosome segregation.The Journal of Cell Biology Sep 2022Kinetochores are multiprotein assemblies directing mitotic spindle attachment and chromosome segregation. In apicomplexan parasites, most known kinetochore components...
Kinetochores are multiprotein assemblies directing mitotic spindle attachment and chromosome segregation. In apicomplexan parasites, most known kinetochore components and associated regulators are apparently missing, suggesting a minimal structure with limited control over chromosome segregation. In this study, we use interactomics combined with deep homology searches to identify 13 previously unknown components of kinetochores in Apicomplexa. Apicomplexan kinetochores are highly divergent in sequence and composition from animal and fungal models. The nanoscale organization includes at least four discrete compartments, each displaying different biochemical interactions, subkinetochore localizations and evolutionary rates across the phylum. We reveal alignment of kinetochores at the metaphase plate in both Plasmodium berghei and Toxoplasma gondii, suggestive of a conserved "hold signal" that prevents precocious entry into anaphase. Finally, we show unexpected plasticity in kinetochore composition and segregation between apicomplexan lifecycle stages, suggestive of diverse requirements to maintain fidelity of chromosome segregation across parasite modes of division.
Topics: Anaphase; Apicomplexa; Chromosome Segregation; Kinetochores; Metaphase; Microtubules; Mitosis; Plasmodium berghei; Spindle Apparatus; Toxoplasma
PubMed: 36006241
DOI: 10.1083/jcb.202111084 -
Proceedings of the National Academy of... Mar 2018Cerebral malaria (CM) is a severe and rapidly progressing complication of infection by parasites that is associated with high rates of mortality and morbidity....
Cerebral malaria (CM) is a severe and rapidly progressing complication of infection by parasites that is associated with high rates of mortality and morbidity. Treatment options are currently few, and intervention with artemisinin (Art) has limited efficacy, a problem that is compounded by the emergence of resistance to Art in parasites. Rocaglates are a class of natural products derived from plants of the genus that have been shown to interfere with eukaryotic initiation factor 4A (eIF4A), ultimately blocking initiation of protein synthesis. Here, we show that the rocaglate CR-1-31B perturbs association of eIF4A (PfeIF4A) with RNA. CR-1-31B shows potent prophylactic and therapeutic antiplasmodial activity in vivo in mouse models of infection with (CM) and (blood-stage malaria), and can also block replication of different clinical isolates of in human erythrocytes infected ex vivo including drug-resistant isolates. In vivo, a single dosing of CR-1-31B in -infected animals is sufficient to provide protection against lethality. CR-1-31B is shown to dampen expression of the early proinflammatory response in myeloid cells in vitro and dampens the inflammatory response in vivo in -infected mice. The dual activity of CR-1-31B as an antiplasmodial and as an inhibitor of the inflammatory response in myeloid cells should prove extremely valuable for therapeutic intervention in human cases of CM.
Topics: Aglaia; Animals; Antimalarials; Disease Models, Animal; Erythrocytes; Eukaryotic Initiation Factor-4F; Female; Humans; Malaria, Cerebral; Mice; Mice, Inbred C57BL; Plant Extracts; Plasmodium berghei; Plasmodium falciparum; Protozoan Proteins
PubMed: 29463745
DOI: 10.1073/pnas.1713000115 -
Scientific Reports Oct 2018Malaria parasite genomes have a range of codon biases, with Plasmodium falciparum one of the most AT-biased genomes known. We examined the make up of synonymous coding... (Comparative Study)
Comparative Study
Malaria parasite genomes have a range of codon biases, with Plasmodium falciparum one of the most AT-biased genomes known. We examined the make up of synonymous coding sites and stop codons in the core genomes of representative malaria parasites, showing first that local DNA context influences codon bias similarly across P. falciparum, P. vivax and P. berghei, with suppression of CpG dinucleotides and enhancement of CpC dinucleotides, both within and aross codons. Intense asexual phase gene expression in P. falciparum and P. berghei is associated with increased A3:G3 bias but reduced T3:C3 bias at 2-fold sites, consistent with adaptation of codons to tRNA pools and avoidance of wobble tRNA interactions that potentially slow down translation. In highly expressed genes, the A3:G3 ratio can exceed 30-fold while the T3:C3 ratio can be less than 1, according to the encoded amino acid and subsequent base. Lysine codons (AAA/G) show distinctive behaviour with substantially reduced A3:G3 bias in highly expressed genes, perhaps because of selection against frameshifting when the AAA codon is followed by another adenine. Intense expression is also associated with a strong bias towards TAA stop codons (found in 94% and 89% of highly expressed P. falciparum and P. berghei genes respectively) and a proportional rise in the TAAA stop 'tetranucleotide'. The presence of these expression-linked effects in the relatively AT-rich malaria parasite species adds weight to the suggestion that AT-richness in the Plasmodium genus might be a fitness adaptation. Potential explanations for the relative lack of codon bias in P. vivax include the distinct features of its lifecycle and its effective population size over evolutionary time.
Topics: Amino Acids; Base Composition; Base Pairing; Codon; DNA, Protozoan; Gene Expression Regulation, Developmental; Genes, Protozoan; Genetic Code; Mutation; Plasmodium berghei; Plasmodium falciparum; Plasmodium vivax; RNA, Protozoan; RNA, Transfer; Selection, Genetic
PubMed: 30374097
DOI: 10.1038/s41598-018-34404-9 -
Malaria Journal Jun 2017Aspartate, which is converted from oxaloacetate (OAA) by aspartate aminotransferase, is considered an important precursor for purine salvage and pyrimidine de novo...
BACKGROUND
Aspartate, which is converted from oxaloacetate (OAA) by aspartate aminotransferase, is considered an important precursor for purine salvage and pyrimidine de novo biosynthesis, and is thus indispensable for the growth of Plasmodium parasites at the asexual blood stages. OAA can be produced in malaria parasites via two routes: (i) from phosphoenolpyruvate (PEP) by phosphoenolpyruvate carboxylase (PEPC) in the cytosol, or (ii) from fumarate by consecutive reactions catalyzed by fumarate hydratase (FH) and malate:quinone oxidoreductase (MQO) in the mitochondria of malaria parasites. Although PEPC-deficient Plasmodium falciparum and Plasmodium berghei (rodent malaria) parasites show a growth defect, the mutant P. berghei can still cause experimental cerebral malaria (ECM) with similar dynamics to wild-type parasites. In contrast, the importance of FH and MQO for parasite viability, growth and virulence is not fully understood because no FH- and MQO-deficient P. falciparum has been established. In this study, the role of FH and MQO in the pathogenicity of asexual-blood-stage Plasmodium parasites causing cerebral malaria was examined.
RESULTS
First, FH- and MQO-deficient parasites were generated by inserting a luciferase-expressing cassette into the fh and mqo loci in the genome of P. berghei ANKA strain. Second, the viability of FH-deficient and MQO-deficient parasites that express luciferase was determined by measuring luciferase activity, and the effect of FH or MQO deficiency on the development of ECM was examined. While the viability of FH-deficient P. berghei was comparable to that of control parasites, MQO-deficient parasites exhibited considerably reduced viability. FH activity derived from erythrocytes was also detected. This result and the absence of phenotype in FH-deficient P. berghei parasites suggest that fumarate can be metabolized to malate by host or parasite FH in P. berghei-infected erythrocytes. Furthermore, although the growth of FH- and MQO-deficient parasites was impaired, the development of ECM was suppressed only in mice infected with MQO-deficient parasites.
CONCLUSIONS
These findings suggest that MQO-mediated mitochondrial functions are required for development of ECM of asexual-blood-stage Plasmodium parasites.
Topics: Animals; Blood-Brain Barrier; Erythrocytes; Female; Fumarate Hydratase; Fumarates; Malaria, Cerebral; Malates; Male; Membrane Potential, Mitochondrial; Mice; Mice, Inbred C57BL; Mitochondria; Oxaloacetic Acid; Oxidoreductases; Plasmodium berghei; Specific Pathogen-Free Organisms
PubMed: 28606087
DOI: 10.1186/s12936-017-1898-5 -
Molecular and Biochemical Parasitology Jul 2021Malaria parasites exhibit a complex lifecycle, requiring extensive asexual replication in the liver and blood of the vertebrate host, and in the haemocoel of the insect... (Review)
Review
Malaria parasites exhibit a complex lifecycle, requiring extensive asexual replication in the liver and blood of the vertebrate host, and in the haemocoel of the insect vector. Yet, they must also undergo a single round of sexual reproduction, which occurs in the vector's midgut upon uptake of a blood meal. Sexual reproduction is obligate for infection of the vector and thus, is essential for onwards transmission to new hosts. Sex in malaria parasites involves several bottlenecks in parasite number, making the stages involved attractive targets for blocking disease transmission. Malaria parasites have evolved a suite of adaptations ("strategies") to maximise the success of sexual reproduction and transmission, which could undermine transmission-blocking interventions. Yet, understanding parasite strategies may also reveal novel opportunities for such interventions. Here, we outline how evolutionary and ecological theories, developed to explain reproductive strategies in multicellular taxa, can be applied to explain two reproductive strategies (conversion rate and sex ratio) expressed by malaria parasites within the vertebrate host.
Topics: Animals; Biological Coevolution; Culicidae; Erythrocytes; Female; Gametogenesis; Host-Parasite Interactions; Humans; Insect Vectors; Life Cycle Stages; Liver; Malaria; Male; Plasmodium berghei; Plasmodium chabaudi; Plasmodium falciparum; Plasmodium knowlesi; Reproduction, Asexual; Sex Ratio
PubMed: 34023299
DOI: 10.1016/j.molbiopara.2021.111375