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Targeted disruption of py235ebp-1: invasion of erythrocytes by Plasmodium yoelii using an alternative Py235 erythrocyte binding protein.

Ogun SA, Tewari R, Otto TD, Howell SA, Knuepfer E, Cunningham DA, Xu Z, Pain A, Holder AA - PLoS Pathog. (2011)

Bottom Line: The PY01365, PY01185, and PY05995/PY03534 genes are part of a distinct subset of the py235 family.In P. falciparum, the RH protein genes are under epigenetic control and expression correlates with binding to distinct erythrocyte receptors and specific invasion pathways, whereas in P. yoelii YM all the genes are expressed and deletion of one does not result in upregulation of another.We propose that simultaneous expression of multiple Py235 ligands enables invasion of a wide range of host erythrocytes even in the presence of antibodies to one or more of the proteins and that this functional redundancy at the protein level gives the parasite phenotypic plasticity in the absence of differences in gene expression.

View Article: PubMed Central - PubMed

Affiliation: Division of Parasitology, MRC National Institute for Medical Research, London, UK. sogun@nimr.mrc.ac.uk

ABSTRACT
Plasmodium yoelii YM asexual blood stage parasites express multiple members of the py235 gene family, part of the super-family of genes including those coding for Plasmodium vivax reticulocyte binding proteins and Plasmodium falciparum RH proteins. We previously identified a Py235 erythrocyte binding protein (Py235EBP-1, encoded by the PY01365 gene) that is recognized by protective mAb 25.77. Proteins recognized by a second protective mAb 25.37 have been identified by mass spectrometry and are encoded by two genes, PY01185 and PY05995/PY03534. We deleted the PY01365 gene and examined the phenotype. The expression of the members of the py235 family in both the WT and gene deletion parasites was measured by quantitative RT-PCR and RNA-Seq. py235ebp-1 expression was undetectable in the knockout parasite, but transcription of other members of the family was essentially unaffected. The knockout parasites continued to react with mAb 25.77; and the 25.77-binding proteins in these parasites were the PY01185 and PY05995/PY03534 products. The PY01185 product was also identified as erythrocyte binding. There was no clear change in erythrocyte invasion profile suggesting that the PY01185 gene product (designated PY235EBP-2) is able to fulfill the role of EBP-1 by serving as an invasion ligand although the molecular details of its interaction with erythrocytes have not been examined. The PY01365, PY01185, and PY05995/PY03534 genes are part of a distinct subset of the py235 family. In P. falciparum, the RH protein genes are under epigenetic control and expression correlates with binding to distinct erythrocyte receptors and specific invasion pathways, whereas in P. yoelii YM all the genes are expressed and deletion of one does not result in upregulation of another. We propose that simultaneous expression of multiple Py235 ligands enables invasion of a wide range of host erythrocytes even in the presence of antibodies to one or more of the proteins and that this functional redundancy at the protein level gives the parasite phenotypic plasticity in the absence of differences in gene expression.

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Determination of the number of copies of PY01365 in the                            genome.Southern blot analysis of WT genomic DNA digested with restriction                            enzymes. Panel A probed with Fragment C and digested with Hinc II and                            Hind III (lane 1), Hinc II and Acc 651 (lane 2), Hinc II and Bgl II                            (lane 3), Hinc II and Pvu I (lane 4) and Hinc II and Sca I (lane 5).                            Panel B probed with Fragment B and digested with Nco I and Sca I (lane                            1), Nco I and Pvu I (lane 2), Nco I and Pst I (lane 3), Nco I and Bgl II                            (lane 4) and Nco I and Acc 651 (lane 5). The migration of size markers                            (Kb) is indicated.
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ppat-1001288-g002: Determination of the number of copies of PY01365 in the genome.Southern blot analysis of WT genomic DNA digested with restriction enzymes. Panel A probed with Fragment C and digested with Hinc II and Hind III (lane 1), Hinc II and Acc 651 (lane 2), Hinc II and Bgl II (lane 3), Hinc II and Pvu I (lane 4) and Hinc II and Sca I (lane 5). Panel B probed with Fragment B and digested with Nco I and Sca I (lane 1), Nco I and Pvu I (lane 2), Nco I and Pst I (lane 3), Nco I and Bgl II (lane 4) and Nco I and Acc 651 (lane 5). The migration of size markers (Kb) is indicated.

Mentions: Southern blot analysis of ten sets of double restriction enzyme digested gDNA from WT P. yoelii YM-parasitized erythrocytes probed with either Fragment B or C gave single bands under low stringency washes (Figure 2). These data suggest that PY01365 is a single copy gene in the line of P. yoelii YM parasites used in this study. This conclusion is also supported by the absence of any RNA-Seq reads mapping to any part of PY01365 from the PY01365-KO parasite.


Targeted disruption of py235ebp-1: invasion of erythrocytes by Plasmodium yoelii using an alternative Py235 erythrocyte binding protein.

Ogun SA, Tewari R, Otto TD, Howell SA, Knuepfer E, Cunningham DA, Xu Z, Pain A, Holder AA - PLoS Pathog. (2011)

Determination of the number of copies of PY01365 in the                            genome.Southern blot analysis of WT genomic DNA digested with restriction                            enzymes. Panel A probed with Fragment C and digested with Hinc II and                            Hind III (lane 1), Hinc II and Acc 651 (lane 2), Hinc II and Bgl II                            (lane 3), Hinc II and Pvu I (lane 4) and Hinc II and Sca I (lane 5).                            Panel B probed with Fragment B and digested with Nco I and Sca I (lane                            1), Nco I and Pvu I (lane 2), Nco I and Pst I (lane 3), Nco I and Bgl II                            (lane 4) and Nco I and Acc 651 (lane 5). The migration of size markers                            (Kb) is indicated.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3040676&req=5

ppat-1001288-g002: Determination of the number of copies of PY01365 in the genome.Southern blot analysis of WT genomic DNA digested with restriction enzymes. Panel A probed with Fragment C and digested with Hinc II and Hind III (lane 1), Hinc II and Acc 651 (lane 2), Hinc II and Bgl II (lane 3), Hinc II and Pvu I (lane 4) and Hinc II and Sca I (lane 5). Panel B probed with Fragment B and digested with Nco I and Sca I (lane 1), Nco I and Pvu I (lane 2), Nco I and Pst I (lane 3), Nco I and Bgl II (lane 4) and Nco I and Acc 651 (lane 5). The migration of size markers (Kb) is indicated.
Mentions: Southern blot analysis of ten sets of double restriction enzyme digested gDNA from WT P. yoelii YM-parasitized erythrocytes probed with either Fragment B or C gave single bands under low stringency washes (Figure 2). These data suggest that PY01365 is a single copy gene in the line of P. yoelii YM parasites used in this study. This conclusion is also supported by the absence of any RNA-Seq reads mapping to any part of PY01365 from the PY01365-KO parasite.

Bottom Line: The PY01365, PY01185, and PY05995/PY03534 genes are part of a distinct subset of the py235 family.In P. falciparum, the RH protein genes are under epigenetic control and expression correlates with binding to distinct erythrocyte receptors and specific invasion pathways, whereas in P. yoelii YM all the genes are expressed and deletion of one does not result in upregulation of another.We propose that simultaneous expression of multiple Py235 ligands enables invasion of a wide range of host erythrocytes even in the presence of antibodies to one or more of the proteins and that this functional redundancy at the protein level gives the parasite phenotypic plasticity in the absence of differences in gene expression.

View Article: PubMed Central - PubMed

Affiliation: Division of Parasitology, MRC National Institute for Medical Research, London, UK. sogun@nimr.mrc.ac.uk

ABSTRACT
Plasmodium yoelii YM asexual blood stage parasites express multiple members of the py235 gene family, part of the super-family of genes including those coding for Plasmodium vivax reticulocyte binding proteins and Plasmodium falciparum RH proteins. We previously identified a Py235 erythrocyte binding protein (Py235EBP-1, encoded by the PY01365 gene) that is recognized by protective mAb 25.77. Proteins recognized by a second protective mAb 25.37 have been identified by mass spectrometry and are encoded by two genes, PY01185 and PY05995/PY03534. We deleted the PY01365 gene and examined the phenotype. The expression of the members of the py235 family in both the WT and gene deletion parasites was measured by quantitative RT-PCR and RNA-Seq. py235ebp-1 expression was undetectable in the knockout parasite, but transcription of other members of the family was essentially unaffected. The knockout parasites continued to react with mAb 25.77; and the 25.77-binding proteins in these parasites were the PY01185 and PY05995/PY03534 products. The PY01185 product was also identified as erythrocyte binding. There was no clear change in erythrocyte invasion profile suggesting that the PY01185 gene product (designated PY235EBP-2) is able to fulfill the role of EBP-1 by serving as an invasion ligand although the molecular details of its interaction with erythrocytes have not been examined. The PY01365, PY01185, and PY05995/PY03534 genes are part of a distinct subset of the py235 family. In P. falciparum, the RH protein genes are under epigenetic control and expression correlates with binding to distinct erythrocyte receptors and specific invasion pathways, whereas in P. yoelii YM all the genes are expressed and deletion of one does not result in upregulation of another. We propose that simultaneous expression of multiple Py235 ligands enables invasion of a wide range of host erythrocytes even in the presence of antibodies to one or more of the proteins and that this functional redundancy at the protein level gives the parasite phenotypic plasticity in the absence of differences in gene expression.

Show MeSH
Related in: MedlinePlus