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A Unique Protein Phosphatase with Kelch-Like Domains (PPKL) in Plasmodium Modulates Ookinete Differentiation, Motility and Invasion

Protein phosphorylation and dephosphorylation (catalysed by kinases and phosphatases, respectively) are post-translational modifications that play key roles in many eukaryotic signalling pathways, and are often deregulated in a number of pathological conditions in humans. In the malaria parasite Pla...

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Autores principales: Guttery, David S., Poulin, Benoit, Ferguson, David J. P., Szöőr, Balázs, Wickstead, Bill, Carroll, Paula L., Ramakrishnan, Chandra, Brady, Declan, Patzewitz, Eva-Maria, Straschil, Ursula, Solyakov, Lev, Green, Judith L., Sinden, Robert E., Tobin, Andrew B., Holder, Anthony A., Tewari, Rita
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2012
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3447748/
https://www.ncbi.nlm.nih.gov/pubmed/23028336
http://dx.doi.org/10.1371/journal.ppat.1002948
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author Guttery, David S.
Poulin, Benoit
Ferguson, David J. P.
Szöőr, Balázs
Wickstead, Bill
Carroll, Paula L.
Ramakrishnan, Chandra
Brady, Declan
Patzewitz, Eva-Maria
Straschil, Ursula
Solyakov, Lev
Green, Judith L.
Sinden, Robert E.
Tobin, Andrew B.
Holder, Anthony A.
Tewari, Rita
author_facet Guttery, David S.
Poulin, Benoit
Ferguson, David J. P.
Szöőr, Balázs
Wickstead, Bill
Carroll, Paula L.
Ramakrishnan, Chandra
Brady, Declan
Patzewitz, Eva-Maria
Straschil, Ursula
Solyakov, Lev
Green, Judith L.
Sinden, Robert E.
Tobin, Andrew B.
Holder, Anthony A.
Tewari, Rita
author_sort Guttery, David S.
collection PubMed
description Protein phosphorylation and dephosphorylation (catalysed by kinases and phosphatases, respectively) are post-translational modifications that play key roles in many eukaryotic signalling pathways, and are often deregulated in a number of pathological conditions in humans. In the malaria parasite Plasmodium, functional insights into its kinome have only recently been achieved, with over half being essential for blood stage development and another 14 kinases being essential for sexual development and mosquito transmission. However, functions for any of the plasmodial protein phosphatases are unknown. Here, we use reverse genetics in the rodent malaria model, Plasmodium berghei, to examine the role of a unique protein phosphatase containing kelch-like domains (termed PPKL) from a family related to Arabidopsis BSU1. Phylogenetic analysis confirmed that the family of BSU1-like proteins including PPKL is encoded in the genomes of land plants, green algae and alveolates, but not in other eukaryotic lineages. Furthermore, PPKL was observed in a distinct family, separate to the most closely-related phosphatase family, PP1. In our genetic approach, C-terminal GFP fusion with PPKL showed an active protein phosphatase preferentially expressed in female gametocytes and ookinetes. Deletion of the endogenous ppkl gene caused abnormal ookinete development and differentiation, and dissociated apical microtubules from the inner-membrane complex, generating an immotile phenotype and failure to invade the mosquito mid-gut epithelium. These observations were substantiated by changes in localisation of cytoskeletal tubulin and actin, and the micronemal protein CTRP in the knockout mutant as assessed by indirect immunofluorescence. Finally, increased mRNA expression of dozi, a RNA helicase vital to zygote development was observed in ppkl(−) mutants, with global phosphorylation studies of ookinete differentiation from 1.5–24 h post-fertilisation indicating major changes in the first hours of zygote development. Our work demonstrates a stage-specific essentiality of the unique PPKL enzyme, which modulates parasite differentiation, motility and transmission.
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spelling pubmed-34477482012-10-01 A Unique Protein Phosphatase with Kelch-Like Domains (PPKL) in Plasmodium Modulates Ookinete Differentiation, Motility and Invasion Guttery, David S. Poulin, Benoit Ferguson, David J. P. Szöőr, Balázs Wickstead, Bill Carroll, Paula L. Ramakrishnan, Chandra Brady, Declan Patzewitz, Eva-Maria Straschil, Ursula Solyakov, Lev Green, Judith L. Sinden, Robert E. Tobin, Andrew B. Holder, Anthony A. Tewari, Rita PLoS Pathog Research Article Protein phosphorylation and dephosphorylation (catalysed by kinases and phosphatases, respectively) are post-translational modifications that play key roles in many eukaryotic signalling pathways, and are often deregulated in a number of pathological conditions in humans. In the malaria parasite Plasmodium, functional insights into its kinome have only recently been achieved, with over half being essential for blood stage development and another 14 kinases being essential for sexual development and mosquito transmission. However, functions for any of the plasmodial protein phosphatases are unknown. Here, we use reverse genetics in the rodent malaria model, Plasmodium berghei, to examine the role of a unique protein phosphatase containing kelch-like domains (termed PPKL) from a family related to Arabidopsis BSU1. Phylogenetic analysis confirmed that the family of BSU1-like proteins including PPKL is encoded in the genomes of land plants, green algae and alveolates, but not in other eukaryotic lineages. Furthermore, PPKL was observed in a distinct family, separate to the most closely-related phosphatase family, PP1. In our genetic approach, C-terminal GFP fusion with PPKL showed an active protein phosphatase preferentially expressed in female gametocytes and ookinetes. Deletion of the endogenous ppkl gene caused abnormal ookinete development and differentiation, and dissociated apical microtubules from the inner-membrane complex, generating an immotile phenotype and failure to invade the mosquito mid-gut epithelium. These observations were substantiated by changes in localisation of cytoskeletal tubulin and actin, and the micronemal protein CTRP in the knockout mutant as assessed by indirect immunofluorescence. Finally, increased mRNA expression of dozi, a RNA helicase vital to zygote development was observed in ppkl(−) mutants, with global phosphorylation studies of ookinete differentiation from 1.5–24 h post-fertilisation indicating major changes in the first hours of zygote development. Our work demonstrates a stage-specific essentiality of the unique PPKL enzyme, which modulates parasite differentiation, motility and transmission. Public Library of Science 2012-09-20 /pmc/articles/PMC3447748/ /pubmed/23028336 http://dx.doi.org/10.1371/journal.ppat.1002948 Text en © 2012 Guttery et al http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.
spellingShingle Research Article
Guttery, David S.
Poulin, Benoit
Ferguson, David J. P.
Szöőr, Balázs
Wickstead, Bill
Carroll, Paula L.
Ramakrishnan, Chandra
Brady, Declan
Patzewitz, Eva-Maria
Straschil, Ursula
Solyakov, Lev
Green, Judith L.
Sinden, Robert E.
Tobin, Andrew B.
Holder, Anthony A.
Tewari, Rita
A Unique Protein Phosphatase with Kelch-Like Domains (PPKL) in Plasmodium Modulates Ookinete Differentiation, Motility and Invasion
title A Unique Protein Phosphatase with Kelch-Like Domains (PPKL) in Plasmodium Modulates Ookinete Differentiation, Motility and Invasion
title_full A Unique Protein Phosphatase with Kelch-Like Domains (PPKL) in Plasmodium Modulates Ookinete Differentiation, Motility and Invasion
title_fullStr A Unique Protein Phosphatase with Kelch-Like Domains (PPKL) in Plasmodium Modulates Ookinete Differentiation, Motility and Invasion
title_full_unstemmed A Unique Protein Phosphatase with Kelch-Like Domains (PPKL) in Plasmodium Modulates Ookinete Differentiation, Motility and Invasion
title_short A Unique Protein Phosphatase with Kelch-Like Domains (PPKL) in Plasmodium Modulates Ookinete Differentiation, Motility and Invasion
title_sort unique protein phosphatase with kelch-like domains (ppkl) in plasmodium modulates ookinete differentiation, motility and invasion
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3447748/
https://www.ncbi.nlm.nih.gov/pubmed/23028336
http://dx.doi.org/10.1371/journal.ppat.1002948
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