9 results on '"Kenthirapalan S"'
Search Results
2. Vital and dispensable roles of Plasmodium multidrug resistance transporters during blood- and mosquito-stage development
- Author
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Rijpma, S.R., Velden, M. van der, Annoura, T., Matz, J.M., Kenthirapalan, S., Kooij, T.W.A., Matuschewski, K., Gemert, G.J. van, Vegte-Bolmer, M. van de, Siebelink-Stoter, R., Graumans, W., Ramesar, J., Klop, O., Russel, F.G.M., Sauerwein, R.W., Janse, C.J., Franke-Fayard, B.M., and Koenderink, J.B.
- Subjects
Renal disorders Radboud Institute for Molecular Life Sciences [Radboudumc 11] ,lnfectious Diseases and Global Health Radboud Institute for Health Sciences [Radboudumc 4] ,parasitic diseases ,lnfectious Diseases and Global Health Radboud Institute for Molecular Life Sciences [Radboudumc 4] - Abstract
Contains fulltext : 170840.pdf (Publisher’s version ) (Closed access) Multidrug resistance (MDR) proteins belong to the B subfamily of the ATP Binding Cassette (ABC) transporters, which export a wide range of compounds including pharmaceuticals. In this study, we used reverse genetics to study the role of all seven Plasmodium MDR proteins during the life cycle of malaria parasites. Four P. berghei genes (encoding MDR1, 4, 6 and 7) were refractory to deletion, indicating a vital role during blood stage multiplication and validating them as potential targets for antimalarial drugs. Mutants lacking expression of MDR2, MDR3 and MDR5 were generated in both P. berghei and P. falciparum, indicating a dispensable role for blood stage development. Whereas P. berghei mutants lacking MDR3 and MDR5 had a reduced blood stage multiplication in vivo, blood stage growth of P. falciparum mutants in vitro was not significantly different. Oocyst maturation and sporozoite formation in Plasmodium mutants lacking MDR2 or MDR5 was reduced. Sporozoites of these P. berghei mutants were capable of infecting mice and life cycle completion, indicating the absence of vital roles during liver stage development. Our results demonstrate vital and dispensable roles of MDR proteins during blood stages and an important function in sporogony for MDR2 and MDR5 in both Plasmodium species.
- Published
- 2016
3. Copper-transporting ATPase is important for malaria parasite fertility
- Author
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Kenthirapalan, S., Waters, A.P., Matuschewski, K., and Kooij, T.W.A.
- Subjects
parasitic diseases ,lnfectious Diseases and Global Health Radboud Institute for Molecular Life Sciences [Radboudumc 4] - Abstract
Item does not contain fulltext Homeostasis of the trace element copper is essential to all eukaryotic life. Copper serves as a cofactor in metalloenzymes and catalyses electron transfer reactions as well as the generation of potentially toxic reactive oxygen species. Here, we describe the functional characterization of an evolutionarily highly conserved, predicted copper-transporting P-type ATPase (CuTP) in the murine malaria model parasite Plasmodium berghei. Live imaging of a parasite line expressing a fluorescently tagged CuTP demonstrated that CuTP is predominantly located in vesicular bodies of the parasite. A P. berghei loss-of-function mutant line was readily obtained and showed no apparent defect in in vivo blood stage growth. Parasite transmission through the mosquito vector was severely affected, but not entirely abolished. We show that male and female gametocytes are abundant in cutp(-) parasites, but activation of male microgametes and exflagellation were strongly impaired. This specific defect could be mimicked by addition of the copper chelator neocuproine to wild-type gametocytes. A cross-fertilization assay demonstrated that female fertility was also severely abrogated. In conclusion, we provide experimental genetic and pharmacological evidence that a healthy copper homeostasis is critical to malaria parasite fertility of both genders of gametocyte and, hence, to transmission to the mosquito vector.
- Published
- 2014
4. Distinct adaptations of a gametocyte ABC transporter to murine and human Plasmodium parasites and its incompatibility in cross-species complementation.
- Author
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Kenthirapalan S, Tran PN, Kooij TWA, Ridgway MC, Rauch M, Brown SHJ, Mitchell TW, Matuschewski K, and Maier AG
- Subjects
- Animals, Female, Humans, Malaria, Falciparum, Mice, ATP-Binding Cassette Transporters genetics, Plasmodium berghei genetics, Plasmodium falciparum genetics, Protozoan Proteins genetics
- Abstract
Parasites of the genus Plasmodium infect a wide range of mammalian hosts including humans, primates, bats and arboreal rodents. A hallmark of Plasmodium spp. is the very narrow host range, indicative of matching parasite-host coevolution. Accordingly, their respective genomes harbour many unique genes and gene families that typically encode proteins involved in host cell recognition and remodelling. Whether and to what extent conserved proteins that are shared across Plasmodium spp. also exert distinct species-specific roles remains largely untested. Here, we present detailed functional profiling of the female gametocyte-specific ATP-binding cassette transporter gABCG2 in the murine parasite Plasmodium berghei and compare our findings with data from the orthologous gene in the human parasite Plasmodium falciparum. We show that P. berghei gABCG2 is female-specific and continues to be expressed in zygotes and ookinetes. In contrast to a distinct localization to Iipid-rich gametocyte-specific spots as observed in P. falciparum, the murine malaria parasite homolog is found at the parasite plasma membrane. Plasmodium berghei lacking gABCG2 displays fast asexual blood-stage replication and increased proportions of female gametocytes, consistent with the corresponding P. falciparum knock-out phenotype. Strikingly, cross-species replacement of gABCG2 in either the murine or the human parasite did not restore normal growth rates. The lack of successful complementation despite high conservation across Plasmodium spp. is an indicator of distinct adaptations and tight parasite-host coevolution. Hence, incompatibility of conserved genes in closely related Plasmodium spp. might be more common than previously anticipated., (Copyright © 2020 Australian Society for Parasitology. Published by Elsevier Ltd. All rights reserved.)
- Published
- 2020
- Full Text
- View/download PDF
5. Vital and dispensable roles of Plasmodium multidrug resistance transporters during blood- and mosquito-stage development.
- Author
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Rijpma SR, van der Velden M, Annoura T, Matz JM, Kenthirapalan S, Kooij TW, Matuschewski K, van Gemert GJ, van de Vegte-Bolmer M, Siebelink-Stoter R, Graumans W, Ramesar J, Klop O, Russel FG, Sauerwein RW, Janse CJ, Franke-Fayard BM, and Koenderink JB
- Subjects
- Animals, Antimalarials pharmacology, Drug Resistance, Multiple, Female, Life Cycle Stages, Malaria parasitology, Malaria, Falciparum parasitology, Male, Membrane Transport Proteins metabolism, Mice, Mice, Inbred BALB C, Mice, Inbred C57BL, Multidrug Resistance-Associated Proteins genetics, Oocytes metabolism, Plasmodium berghei genetics, Plasmodium berghei growth & development, Plasmodium falciparum genetics, Plasmodium falciparum growth & development, Sporozoites metabolism, Culicidae parasitology, Multidrug Resistance-Associated Proteins metabolism, Plasmodium berghei drug effects, Plasmodium berghei metabolism, Plasmodium falciparum drug effects, Plasmodium falciparum metabolism
- Abstract
Multidrug resistance (MDR) proteins belong to the B subfamily of the ATP Binding Cassette (ABC) transporters, which export a wide range of compounds including pharmaceuticals. In this study, we used reverse genetics to study the role of all seven Plasmodium MDR proteins during the life cycle of malaria parasites. Four P. berghei genes (encoding MDR1, 4, 6 and 7) were refractory to deletion, indicating a vital role during blood stage multiplication and validating them as potential targets for antimalarial drugs. Mutants lacking expression of MDR2, MDR3 and MDR5 were generated in both P. berghei and P. falciparum, indicating a dispensable role for blood stage development. Whereas P. berghei mutants lacking MDR3 and MDR5 had a reduced blood stage multiplication in vivo, blood stage growth of P. falciparum mutants in vitro was not significantly different. Oocyst maturation and sporozoite formation in Plasmodium mutants lacking MDR2 or MDR5 was reduced. Sporozoites of these P. berghei mutants were capable of infecting mice and life cycle completion, indicating the absence of vital roles during liver stage development. Our results demonstrate vital and dispensable roles of MDR proteins during blood stages and an important function in sporogony for MDR2 and MDR5 in both Plasmodium species., (© 2016 John Wiley & Sons Ltd.)
- Published
- 2016
- Full Text
- View/download PDF
6. Functional profiles of orphan membrane transporters in the life cycle of the malaria parasite.
- Author
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Kenthirapalan S, Waters AP, Matuschewski K, and Kooij TW
- Subjects
- Animals, Anopheles, Female, Life Cycle Stages, Male, Membrane Transport Proteins genetics, Mice, Mice, Inbred C57BL, Plasmodium berghei genetics, Membrane Transport Proteins metabolism, Plasmodium berghei metabolism
- Abstract
Assigning function to orphan membrane transport proteins and prioritizing candidates for detailed biochemical characterization remain fundamental challenges and are particularly important for medically relevant pathogens, such as malaria parasites. Here we present a comprehensive genetic analysis of 35 orphan transport proteins of Plasmodium berghei during its life cycle in mice and Anopheles mosquitoes. Six genes, including four candidate aminophospholipid transporters, are refractory to gene deletion, indicative of essential functions. We generate and phenotypically characterize 29 mutant strains with deletions of individual transporter genes. Whereas seven genes appear to be dispensable under the experimental conditions tested, deletion of any of the 22 other genes leads to specific defects in life cycle progression in vivo and/or host transition. Our study provides growing support for a potential link between heavy metal homeostasis and host switching and reveals potential targets for rational design of new intervention strategies against malaria.
- Published
- 2016
- Full Text
- View/download PDF
7. Copper-transporting ATPase is important for malaria parasite fertility.
- Author
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Kenthirapalan S, Waters AP, Matuschewski K, and Kooij TW
- Subjects
- Animals, Copper-Transporting ATPases, Disease Models, Animal, Female, Fertility, Malaria metabolism, Male, Mice, Mice, Inbred C57BL, Mutation, Phenanthrolines pharmacology, Plasmodium berghei genetics, Plasmodium berghei growth & development, Plasmodium berghei pathogenicity, Adenosine Triphosphatases metabolism, Cation Transport Proteins metabolism, Copper metabolism, Culicidae parasitology, Malaria parasitology, Plasmodium berghei enzymology, Protozoan Proteins metabolism
- Abstract
Homeostasis of the trace element copper is essential to all eukaryotic life. Copper serves as a cofactor in metalloenzymes and catalyses electron transfer reactions as well as the generation of potentially toxic reactive oxygen species. Here, we describe the functional characterization of an evolutionarily highly conserved, predicted copper-transporting P-type ATPase (CuTP) in the murine malaria model parasite Plasmodium berghei. Live imaging of a parasite line expressing a fluorescently tagged CuTP demonstrated that CuTP is predominantly located in vesicular bodies of the parasite. A P. berghei loss-of-function mutant line was readily obtained and showed no apparent defect in in vivo blood stage growth. Parasite transmission through the mosquito vector was severely affected, but not entirely abolished. We show that male and female gametocytes are abundant in cutp(-) parasites, but activation of male microgametes and exflagellation were strongly impaired. This specific defect could be mimicked by addition of the copper chelator neocuproine to wild-type gametocytes. A cross-fertilization assay demonstrated that female fertility was also severely abrogated. In conclusion, we provide experimental genetic and pharmacological evidence that a healthy copper homeostasis is critical to malaria parasite fertility of both genders of gametocyte and, hence, to transmission to the mosquito vector., (© 2013 The Authors. Molecular Microbiology published by John Wiley & Sons Ltd.)
- Published
- 2014
- Full Text
- View/download PDF
8. Flow cytometry-assisted rapid isolation of recombinant Plasmodium berghei parasites exemplified by functional analysis of aquaglyceroporin.
- Author
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Kenthirapalan S, Waters AP, Matuschewski K, and Kooij TW
- Subjects
- Animals, Gene Expression Regulation physiology, Genotype, Mice, Mutation, Organisms, Genetically Modified, Plasmodium berghei metabolism, Aquaglyceroporins chemistry, Aquaglyceroporins metabolism, Flow Cytometry methods, Malaria parasitology, Plasmodium berghei genetics, Plasmodium berghei isolation & purification
- Abstract
The most critical bottleneck in the generation of recombinant Plasmodium berghei parasites is the mandatory in vivo cloning step following successful genetic manipulation. This study describes a new technique for rapid selection of recombinant P. berghei parasites. The method is based on flow cytometry to isolate isogenic parasite lines and represents a major advance for the field, in that it will speed the generation of recombinant parasites as well as cut down on animal use significantly. High expression of GFP during blood infection, a prerequisite for robust separation of transgenic lines by flow cytometry, was achieved. Isogenic recombinant parasite populations were isolated even in the presence of a 100-fold excess of wild-type (WT) parasites. Aquaglyceroporin (AQP) loss-of-function mutants and parasites expressing a tagged AQP were generated to validate this approach. aqp(-) parasites grow normally within the WT phenotypic range during blood infection of NMRI mice. Similarly, colonization of the insect vector and establishment of an infection after mosquito transmission were unaffected, indicating that AQP is dispensable for life cycle progression in vivo under physiological conditions, refuting its use as a suitable drug target. Tagged AQP localized to perinuclear structures and not the parasite plasma membrane. We suggest that flow-cytometric isolation of isogenic parasites overcomes the major roadblock towards a genome-scale repository of mutant and transgenic malaria parasite lines., (Copyright © 2012 Australian Society for Parasitology Inc. Published by Elsevier Ltd. All rights reserved.)
- Published
- 2012
- Full Text
- View/download PDF
9. Reprogramming a module of the 6-deoxyerythronolide B synthase for iterative chain elongation.
- Author
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Kapur S, Lowry B, Yuzawa S, Kenthirapalan S, Chen AY, Cane DE, and Khosla C
- Subjects
- Acyl Carrier Protein metabolism, Amino Acid Sequence, Biocatalysis, Models, Molecular, Molecular Sequence Data, Polyketides chemistry, Protein Engineering, Protein Transport, Substrate Specificity, Polyketide Synthases chemistry, Polyketide Synthases metabolism, Polyketides metabolism
- Abstract
Multimodular polyketide synthases (PKSs) have an assembly line architecture in which a set of protein domains, known as a module, participates in one round of polyketide chain elongation and associated chemical modifications, after which the growing chain is translocated to the next PKS module. The ability to rationally reprogram these assembly lines to enable efficient synthesis of new polyketide antibiotics has been a long-standing goal in natural products biosynthesis. We have identified a ratchet mechanism that can explain the observed unidirectional translocation of the growing polyketide chain along the 6-deoxyerythronolide B synthase. As a test of this model, module 3 of the 6-deoxyerythronolide B synthase has been reengineered to catalyze two successive rounds of chain elongation. Our results suggest that high selectivity has been evolutionarily programmed at three types of protein-protein interfaces that are present repetitively along naturally occurring PKS assembly lines.
- Published
- 2012
- Full Text
- View/download PDF
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