Judith M Pfahler

Universität Heidelberg, Heidelburg, Baden-Württemberg, Germany

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Publications (6)24.09 Total impact

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    ABSTRACT: Here, we describe the methodology for transient transfection of Plasmodium vivax. The ability to genetically manipulate P. vivax has rendered this important human malaria parasite more amenable to molecular investigation. However, a systematic analysis of this parasite and its disease-mediating interactions with the human host still awaits further technological breakthroughs, foremost the establishment of a continuous in vitro culture system. Nevertheless, the first steps towards domesticating P. vivax for research purposes have been made. Transfection will eventually help to better understand the unique pathogenic features displayed by P. vivax, such as the host cell specificity for reticulocytes and the occurrence of relapses. Transfection will also be an invaluable tool for studies related to the emerging drug resistance, and it will help identify and validate novel targets for rational intervention.
    No preview · Article · Jan 2013 · Methods in molecular biology (Clifton, N.J.)
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    ABSTRACT: The human malarial parasite Plasmodium falciparum exports determinants of virulence and pathology to destinations within its host erythrocyte, including the cytoplasm, the plasma membrane and membrane profiles of parasite origin termed Maurer's clefts. While there is some information regarding the signals that allot proteins for export, the trafficking route itself has remained largely obscure, partly due to technical limitations in following protein trafficking with time. To overcome these shortcomings, we have established a conditional protein export system in P. falciparum, based on the previously described conditional aggregation domain (CAD domain) that self-aggregates in the endoplasmic reticulum in a manner that is reversible by the addition of a small molecule. By fusing the CAD domain to the first 80 amino acids of STEVOR and full-length PfSBP1, we were able to control export of a soluble and a transmembrane protein to the erythrocyte cytosol and the Maurer's clefts respectively. The conditional export system allowed us to study the temporal sequence of events of protein export and identify intermediate steps. We further explored the potential of the conditional export system in identifying factors that interact with exported proteins en route. Our data provide evidence for a physical interaction of exported proteins with the molecular chaperone PfBiP during early export steps.
    Preview · Article · Sep 2008 · Cellular Microbiology
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    ABSTRACT: The ability to control expression of a specific gene is a prerequisite to understand the function of essential genes. Many gene regulation systems operate on the transcriptional level by employing heterologous cis- and trans-acting elements. Recently, novel approaches employing autocatalytic RNA have been reported for different organisms. Here we show specific downregulation of gene expression in the apicomplexan parasites Toxoplasma gondii and Plasmodium falciparum, employing self-cleaving ribozymes integrated into the transcriptional unit of different genes. Moreover, we demonstrate the potential to specifically upregulate reporter gene expression by employment of the recently identified ribozyme inhibitor toyocamycin. At the RNA-level, we were able to significantly stabilise the mRNA in T. gondii. Furthermore we show that the adenosine analogue toyocamycin needs to be phosphorylated by adenosine kinase in order to act as an inhibitor for hammerhead ribozymes, since neither upregulation of reporter gene expression nor a toxic effect of toyocamycin can be detected in parasites that do not express the enzyme adenosine kinase.
    Full-text · Article · Jun 2008 · International Journal for Parasitology
  • Elise Schieck · Judith M Pfahler · Cecilia P Sanchez · Michael Lanzer

    No preview · Article · Jul 2007 · Molecular and Biochemical Parasitology
  • Judith M Pfahler · Mary R Galinski · John W Barnwell · Michael Lanzer

    No preview · Article · Oct 2006 · Molecular and Biochemical Parasitology
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    ABSTRACT: The human malarial parasite Plasmodium falciparum exports proteins to destinations within its host erythrocyte, including cytosol, surface and membranous profiles of parasite origin termed Maurer's clefts. Although several of these exported proteins are determinants of pathology and virulence, the mechanisms and trafficking signals underpinning protein export are largely uncharacterized-particularly for exported transmembrane proteins. Here, we have investigated the signals mediating trafficking of STEVOR, a family of transmembrane proteins located at the Maurer's clefts and believed to play a role in antigenic variation. Our data show that, apart from a signal sequence, a minimum of two addition signals are required. This includes a host cell targeting signal for export to the host erythrocyte and a transmembrane domain for final sorting to Maurer's clefts. Biochemical studies indicate that STEVOR traverses the secretory pathway as an integral membrane protein. Our data suggest general principles for transport of transmembrane proteins to the Maurer's clefts and provide new insights into protein sorting and trafficking processes in P. falciparum.
    Full-text · Article · Aug 2005 · The EMBO Journal