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Dive into the research topics where Klaus Lingelbach is active.

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Featured researches published by Klaus Lingelbach.


Molecular Microbiology | 2003

Characterization of the pathway for transport of the cytoadherence-mediating protein, PfEMP1, to the host cell surface in malaria parasite-infected erythrocytes

Neline Kriek; Leann Tilley; Paul Horrocks; Robert Pinches; Barry C. Elford; David J. P. Ferguson; Klaus Lingelbach; Chris Newbold

The Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) family of antigenically diverse proteins is expressed on the surface of human erythrocytes infected with the malaria parasite P. falciparum, and mediates cytoadherence to the host vascular endothelium. In this report, we show that export of PfEMP1 is slow and inefficient as it takes several hours to traffic newly synthesized proteins to the erythrocyte membrane. Upon removal by trypsin treatment, the surface‐exposed population of PfEMP1 is not replenished during subsequent culture indicating that there is no cycling of PfEMP1 between the erythrocyte surface and an intracellular compartment. The role of Maurers clefts as an intermediate sorting compartment in trafficking of PfEMP1 was investigated using immunoelectron microscopy and proteolytic digestion of streptolysin O‐permeabilized parasitized erythrocytes. We show that PfEMP1 is inserted into the Maurers cleft membrane with the C‐terminal domain exposed to the erythrocyte cytoplasm, whereas the N‐terminal domain is buried inside the cleft. Transfer of PfEMP1 to the erythrocyte surface appears to involve electron‐lucent extensions of the Maurers clefts. Thus, we have delineated some important aspects of the unusual trafficking mechanism for delivery of this critical parasite virulence factor to the erythrocyte surface.


Molecular Microbiology | 2004

A potential novel mechanism for the insertion of a membrane protein revealed by a biochemical analysis of the Plasmodium falciparum cytoadherence molecule PfEMP-1.

Janni Papakrivos; Chris Newbold; Klaus Lingelbach

Plasmodium falciparum erythrocyte membrane protein‐1 (PfEMP‐1) is exposed on the surface of infected erythrocytes where it both acts as an important pathogenicity factor in malaria and undergoes antigenic variation as a means of immune evasion. Because the mammalian erythrocyte lacks a protein secretory machinery there has been much interest in elucidating the mechanism whereby this protein is transferred from its site of synthesis within the parasite to its final destination. Current opinion favours a mechanism whereby PfEMP‐1 becomes cotranslationally inserted into the endoplasmic reticulum of the parasite and is subsequently transported as an integral part of an erythrocyte cytoplasmic membrane system derived from the parasite. Here we show that the solubility characteristics of this protein during several stages of its transport pathway are inconsistent with this view. Instead we propose that the protein is synthesized as a peripheral membrane protein which only when it arrives at its final destination assumes a transmembrane topology. Even in this state, the extractability of the protein with urea suggest that it is anchored in the membrane by protein‐protein rather than by protein–lipid interaction.


Molecular Microbiology | 2009

Protein unfolding is an essential requirement for transport across the parasitophorous vacuolar membrane of Plasmodium falciparum

Nina Gehde; Corinna Hinrichs; Irine Montilla; Stefan Charpian; Klaus Lingelbach; Jude M. Przyborski

Plasmodium falciparum traffics a large number of proteins to its host cell, the mature human erythrocyte. How exactly these proteins gain access to the red blood cell is poorly understood. Here we have investigated the effect of protein folding on the transport of model substrate proteins to the host cell. We find that proteins must pass into the erythrocyte cytoplasm in an unfolded state. Our data strongly support the presence of a protein‐conducing channel in the parasitophorous vacoular membrane, and additionally imply an important role for molecular chaperones in keeping parasite proteins in a ‘translocation competent’ state prior to membrane passage.


Cellular Microbiology | 2010

Parasite-encoded Hsp40 proteins define novel mobile structures in the cytosol of the P. falciparum-infected erythrocyte

Simone Külzer; Melanie Rug; Klaus Brinkmann; Ping Cannon; Alan F. Cowman; Klaus Lingelbach; Alexander G. Maier; Jude M. Przyborski

Plasmodium falciparum is predicted to transport over 300 proteins to the cytosol of its chosen host cell, the mature human erythrocyte, including 19 members of the Hsp40 family. Here, we have generated transfectant lines expressing GFP‐ or HA‐Strep‐tagged versions of these proteins, and used these to investigate both localization and other properties of these Hsp40 co‐chaperones. These fusion proteins labelled punctate structures within the infected erythrocyte, initially suggestive of a Maurers clefts localization. Further experiments demonstrated that these structures were distinct from the Maurers clefts in protein composition. Transmission electron microscopy verifies a non‐cleft localization for HA‐Strep‐tagged versions of these proteins. We were not able to label these structures with BODIPY–ceramide, suggesting a lower size and/or different lipid composition compared with the Maurers clefts. Solubility studies revealed that the Hsp40–GFP fusion proteins appear to be tightly associated with membranes, but could be released from the bilayer under conditions affecting membrane cholesterol content or organization, suggesting interaction with a binding partner localized to cholesterol‐rich domains. These novel structures are highly mobile in the infected erythrocyte, but based on velocity calculations, can be distinguished from the ‘highly mobile vesicles’ previously described. Our study identifies a further extra‐parasitic structure in the P. falciparum‐infected erythrocyte, which we name ‘J‐dots’ (as their defining characteristic so far is the content of J‐proteins). We suggest that these J‐dots are involved in trafficking of parasite‐encoded proteins through the cytosol of the infected erythrocyte.


PLOS ONE | 2011

Fosmidomycin Uptake into Plasmodium and Babesia-Infected Erythrocytes Is Facilitated by Parasite-Induced New Permeability Pathways

Stefan Baumeister; Jochen Wiesner; Armin Reichenberg; Martin Hintz; Sven Bietz; Omar S. Harb; David S. Roos; Maximilian Kordes; Johannes Friesen; Klaus Lingelbach; Hassan Jomaa; Frank Seeber

Background Highly charged compounds typically suffer from low membrane permeability and thus are generally regarded as sub-optimal drug candidates. Nonetheless, the highly charged drug fosmidomycin and its more active methyl-derivative FR900098 have proven parasiticidal activity against erythrocytic stages of the malaria parasite Plasmodium falciparum. Both compounds target the isoprenoid biosynthesis pathway present in bacteria and plastid-bearing organisms, like apicomplexan parasites. Surprisingly, the compounds are inactive against a range of apicomplexans replicating in nucleated cells, including Toxoplasma gondii. Methodology/Principal Findings Since non-infected erythrocytes are impermeable for FR90098, we hypothesized that these drugs are taken up only by erythrocytes infected with Plasmodium. We provide evidence that radiolabeled FR900098 accumulates in theses cells as a consequence of parasite-induced new properties of the host cell, which coincide with an increased permeability of the erythrocyte membrane. Babesia divergens, a related parasite that also infects human erythrocytes and is also known to induce an increase in membrane permeability, displays a similar susceptibility and uptake behavior with regard to the drug. In contrast, Toxoplasma gondii-infected cells do apparently not take up the compounds, and the drugs are inactive against the liver stages of Plasmodium berghei, a mouse malaria parasite. Conclusions/Significance Our findings provide an explanation for the observed differences in activity of fosmidomycin and FR900098 against different Apicomplexa. These results have important implications for future screens aimed at finding new and safe molecular entities active against P. falciparum and related parasites. Our data provide further evidence that parasite-induced new permeability pathways may be exploited as routes for drug delivery.


Molecular Microbiology | 2006

Evidence for the involvement of Plasmodium falciparum proteins in the formation of new permeability pathways in the erythrocyte membrane

Stefan Baumeister; Markus Winterberg; Christophe Duranton; Stephan M. Huber; Florian Lang; Kiaran Kirk; Klaus Lingelbach

The intraerythrocytic developmental stages of the malaria parasite Plasmodium falciparum are responsible for the clinical symptoms associated with malaria tropica. The non‐infected human erythrocyte is a terminally differentiated cell that is unable to synthesize proteins and lipids de novo, and it is incapable of importing a number of solutes that are essential for parasite proliferation. Approximately 12–15 h after invasion the parasitized cell undergoes a marked increase in its permeability to a variety of different solutes present in the extracellular milieu. The increase is due to the induction in the erythrocyte membrane of ‘new permeability pathways’ which have been characterized in some detail in terms of their transport and electrophysiological properties, but which are yet to be defined at a molecular level. Here we show that these pathways are resistant to trypsin but are abolished by treatment of intact infected erythrocytes with chymotrypsin. On resuspension of chymotrypsinized cells in chymotrypsin‐free medium the pathways progressively reappear, a process that can be inhibited by cytotoxic agents, and by brefeldin A which inhibits protein secretion. Our results provide evidence for the involvement of parasite encoded proteins in the generation of the pathways, either as components of the pathways themselves or as auxiliary factors.


Nucleic Acids Research | 2014

The evolutionary dynamics of variant antigen genes in Babesia reveal a history of genomic innovation underlying host-parasite interaction

Andrew P. Jackson; Thomas D. Otto; Alistair C. Darby; Abhinay Ramaprasad; Dong Xia; Ignacio Echaide; Marisa Farber; Sunayna Gahlot; John Gamble; Dinesh Gupta; Yask Gupta; Louise Jackson; Laurence Malandrin; Tareq B. Malas; Ehab Moussa; Mridul Nair; Adam J. Reid; Mandy Sanders; Jyotsna Sharma; Alan Tracey; Michael A. Quail; William Weir; Jonathan M. Wastling; Neil Hall; Peter Willadsen; Klaus Lingelbach; Brian Shiels; Andy Tait; Matthew Berriman; David R. Allred

Babesia spp. are tick-borne, intraerythrocytic hemoparasites that use antigenic variation to resist host immunity, through sequential modification of the parasite-derived variant erythrocyte surface antigen (VESA) expressed on the infected red blood cell surface. We identified the genomic processes driving antigenic diversity in genes encoding VESA (ves1) through comparative analysis within and between three Babesia species, (B. bigemina, B. divergens and B. bovis). Ves1 structure diverges rapidly after speciation, notably through the evolution of shortened forms (ves2) from 5′ ends of canonical ves1 genes. Phylogenetic analyses show that ves1 genes are transposed between loci routinely, whereas ves2 genes are not. Similarly, analysis of sequence mosaicism shows that recombination drives variation in ves1 sequences, but less so for ves2, indicating the adoption of different mechanisms for variation of the two families. Proteomic analysis of the B. bigemina PR isolate shows that two dominant VESA1 proteins are expressed in the population, whereas numerous VESA2 proteins are co-expressed, consistent with differential transcriptional regulation of each family. Hence, VESA2 proteins are abundant and previously unrecognized elements of Babesia biology, with evolutionary dynamics consistently different to those of VESA1, suggesting that their functions are distinct.


Cellular Physiology and Biochemistry | 2008

A high specificity and affinity interaction with serum albumin stimulates an anion conductance in malaria-infected erythrocytes.

Christophe Duranton; Valerie Tanneur; Camelia Lang; Verena B. Brand; Saisudha Koka; Ravi S. Kasinathan; Martina Dorsch; Hans J. Hedrich; Stefan Baumeister; Klaus Lingelbach; Florian Lang; Stephan M. Huber

The intraerythrocytic development of P. falciparum induces New Permeability Pathways (NPP) in the membrane of the parasitized erythrocyte which provide the parasite with nutrients, adjust the erythrocyte electrolyte composition to the needs of the parasite, and dispose of metabolic waste products and osmolytes. Patch-clamp recordings identified inwardly and outwardly rectifying (OR) anion conductances in the host erythrocyte membrane as electrophysiological correlate of the NPP. The OR conductance is regulated by serum. Here we show that serum albumin (SA) stimulated OR-generated Cl- and lactate outward currents with an EC50 of approximately 100 nM while other proteins such as ovalbumin or casein did not. The stimulatory efficacy did not differ between fatty acid free bovine SA and recombinant human SA and disruption of the SA tertiary structure abolished the effect suggesting that intact SA protein and not other bound factors interact with the erythrocyte membrane. Taken together, the data indicate a high affinity and specificity interaction of native SA with the parasitized erythrocytes which might underlie the observed dependence of P. falciparum growth in vitro on SA.


Molecular and Biochemical Parasitology | 1998

CHARACTERIZATION AND CLONING OF THE GENE ENCODING THE VACUOLAR MEMBRANE PROTEIN EXP-2 FROM PLASMODIUM FALCIPARUM

Thomas Marti; Barbara Rick; Delia Johnson; Jürgen Benting; Stefan Baumeister; Claudia Helmbrecht; Michael Lanzer; Klaus Lingelbach

As a contribution to the characterization of the parasitophorous vacuolar membrane from Plasmodium falciparum we have begun the identification of vacuolar membrane proteins. Exported protein-2 (EXP-2) is a vacuolar membrane protein exposed into the vacuolar space. To further characterize EXP-2, it was purified, and the 45 N-terminal amino acids were determined by micro-sequencing. Based on this information, partial cDNA and genomic fragments were amplified by PCR and used as probes for the isolation of complete cDNA and genomic DNA clones. The single copy gene is located on chromosome 14, and is transcribed during the ring stage of parasite development. The open reading frame encodes an N-terminal signal sequence which is cleaved from the mature protein. The amino acid composition of EXP-2 is characterized by charged amino acids, with a high abundance of aspartate residues in the C-terminal portion of the protein. In contrast to EXP-1, an integral protein of the vacuolar membrane, EXP-2 lacks a typical hydrophobic transmembrane domain. We suggest that EXP-2 may associate with the vacuolar membrane via an amphipathic helix located in the N-terminal half of the protein.


Blood | 2012

Digestive vacuole of Plasmodium falciparum released during erythrocyte rupture dually activates complement and coagulation.

Prasad Dasari; Sophia D. Heber; Maike Beisele; Michael Torzewski; Kurt Reifenberg; Carolin Orning; Anja Fries; Anna-Lena Zapf; Stefan Baumeister; Klaus Lingelbach; Rachanee Udomsangpetch; Sebastian Chakrit Bhakdi; Karina Reiss; Sucharit Bhakdi

Severe Plasmodium falciparum malaria evolves through the interplay among capillary sequestration of parasitized erythrocytes, deregulated inflammatory responses, and hemostasis dysfunction. After rupture, each parasitized erythrocyte releases not only infective merozoites, but also the digestive vacuole (DV), a membrane-bounded organelle containing the malaria pigment hemozoin. In the present study, we report that the intact organelle, but not isolated hemozoin, dually activates the alternative complement and the intrinsic clotting pathway. Procoagulant activity is destroyed by phospholipase C treatment, indicating a critical role of phospholipid head groups exposed at the DV surface. Intravenous injection of DVs caused alternative pathway complement consumption and provoked apathy and reduced nociceptive responses in rats. Ultrasonication destroyed complement-activating and procoagulant properties in vitro and rendered the DVs biologically inactive in vivo. Low-molecular-weight dextran sulfate blocked activation of both complement and coagulation and protected animals from the harmful effects of DV infusion. We surmise that in chronic malaria, complement activation by and opsonization of the DV may serve a useful function in directing hemozoin to phagocytic cells for safe disposal. However, when the waste disposal system of the host is overburdened, DVs may transform into a trigger of pathology and therefore represent a potential therapeutic target in severe malaria.

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Julius O. Nyalwidhe

Eastern Virginia Medical School

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Kiaran Kirk

Australian National University

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Markus Winterberg

Australian National University

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Florian Lang

University of Tübingen

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