Semaphorin-7A Is an Erythrocyte Receptor for P. falciparum Merozoite-Specific TRAP Homolog, MTRAP

The motility and invasion of Plasmodium parasites is believed to require a cytoplasmic actin-myosin motor associated with a cell surface ligand belonging to the TRAP (thrombospondin-related anonymous protein) family. Current models of invasion usually invoke the existence of specific receptors for the TRAP-family ligands on the surface of the host cell; however, the identities of these receptors remain largely unknown. Here, we identify the GPI-linked protein Semaphorin-7A (CD108) as an erythrocyte receptor for the P. falciparum merozoite-specific TRAP homolog (MTRAP) by using a systematic screening approach designed to detect extracellular protein interactions. The specificity of the interaction was demonstrated by showing that binding was saturable and by quantifying the equilibrium and kinetic biophysical binding parameters using surface plasmon resonance. We found that two MTRAP monomers interact via their tandem TSR domains with the Sema domains of a Semaphorin-7A homodimer. Known naturally-occurring polymorphisms in Semaphorin-7A did not quantitatively affect MTRAP binding nor did the presence of glycans on the receptor. Attempts to block the interaction during in vitro erythrocyte invasion assays using recombinant proteins and antibodies showed no significant inhibitory effect, suggesting the inaccessibility of the complex to proteinaceous blocking agents. These findings now provide important experimental evidence to support the model that parasite TRAP-family ligands interact with specific host receptors during cellular invasion.

[1]  D. Stuart,et al.  The ligand-binding face of the semaphorins revealed by the high-resolution crystal structure of SEMA4D , 2003, Nature Structural Biology.

[2]  Alex Bateman,et al.  Large-scale screening for novel low-affinity extracellular protein interactions. , 2008, Genome research.

[3]  G. Blatch,et al.  Heterologous expression of plasmodial proteins for structural studies and functional annotation , 2008, Malaria Journal.

[4]  Juraj Kabat,et al.  Glycophorin B is the erythrocyte receptor of Plasmodium falciparum erythrocyte-binding ligand, EBL-1 , 2009, Proceedings of the National Academy of Sciences.

[5]  T. Speed,et al.  Identification of Proteins from Plasmodium falciparum That Are Homologous to Reticulocyte Binding Proteins inPlasmodium vivax , 2001, Infection and Immunity.

[6]  C. H. Bell,et al.  Structural basis of semaphorin–plexin signalling , 2010, Nature.

[7]  D. DeLuca,et al.  The molecular diversity of Sema7A, the semaphorin that carries the JMH blood group antigens , 2007, Transfusion.

[8]  V. Nussenzweig,et al.  Antibodies to Plasmodium circumsporozoite protein (CSP) inhibit sporozoite's cell traversal activity. , 2012, Journal of immunological methods.

[9]  Y. Durocher,et al.  High-level and high-throughput recombinant protein production by transient transfection of suspension-growing human 293-EBNA1 cells. , 2002, Nucleic acids research.

[10]  A. Kolodkin,et al.  Semaphorin 7A initiates T-cell-mediated inflammatory responses through alpha1beta1 integrin. , 2007, Nature.

[11]  J. Cerny,et al.  Characterization of the human leukocyte GPI-anchored glycoprotein CDw108 and its relation to other similar molecules. , 1999, Immunobiology.

[12]  A. Kolodkin,et al.  Semaphorin 7A initiates T-cell-mediated inflammatory responses through α1β1 integrin , 2007, Nature.

[13]  I. Coppens,et al.  Sites of interaction between aldolase and thrombospondin-related anonymous protein in plasmodium. , 2003, Molecular biology of the cell.

[14]  Min Lu,et al.  Structure of the Semaphorin-3A Receptor Binding Module , 2003, Neuron.

[15]  R. Sinden,et al.  Vital functions of the malarial ookinete protein, CTRP, reside in the A domains. , 2011, International journal for parasitology.

[16]  Nancy Fullman,et al.  Global malaria mortality between 1980 and 2010: a systematic analysis , 2012, The Lancet.

[17]  I. Coppens,et al.  Apicomplexan gliding motility and host cell invasion: overhauling the motor model. , 2004, Trends in parasitology.

[18]  Gavin J. Wright,et al.  Jamb and Jamc Are Essential for Vertebrate Myocyte Fusion , 2011, PLoS biology.

[19]  Gavin J. Wright.  Signal initiation in biological systems: the properties and detection of transient extracellular protein interactions† †This article is part of a Molecular BioSystems themed issue on Computational and Systems Biology. , 2009, Molecular bioSystems.

[20]  Pawan Malhotra,et al.  Structural and functional dissection of the adhesive domains of Plasmodium falciparum thrombospondin-related anonymous protein (TRAP). , 2004, The Biochemical journal.

[21]  H. Bujard,et al.  Thrombospondin related anonymous protein (TRAP) of Plasmodium falciparum binds specifically to sulfated glycoconjugates and to HepG2 hepatoma cells suggesting a role for this molecule in sporozoite invasion of hepatocytes. , 1993, The EMBO journal.

[22]  C. Janse,et al.  PTRAMP; a conserved Plasmodium thrombospondin-related apical merozoite protein. , 2004, Molecular and biochemical parasitology.

[23]  M. Poo,et al.  Unified Nomenclature for the Semaphorins/Collapsins , 1999, Cell.

[24]  K. Matuschewski,et al.  Role for the Plasmodium sporozoite-specific transmembrane protein S6 in parasite motility and efficient malaria transmission , 2008, Cellular microbiology.

[25]  Michel Theron,et al.  An Adaptable Two-Color Flow Cytometric Assay to Quantitate the Invasion of Erythrocytes by Plasmodium falciparum Parasites , 2010, Cytometry. Part A : the journal of the International Society for Analytical Cytology.

[26]  J. Haynes,et al.  A malaria invasion receptor, the 175-kilodalton erythrocyte binding antigen of Plasmodium falciparum recognizes the terminal Neu5Ac(alpha 2- 3)Gal- sequences of glycophorin A , 1992, The Journal of cell biology.

[27]  X. Ambroggio,et al.  Analysis of the Conformation and Function of the Plasmodium falciparum Merozoite Proteins MTRAP and PTRAMP , 2012, Eukaryotic Cell.

[28]  K. Robson.  Thrombospondin-related adhesive protein (TRAP) of Plasmodium falciparum: Expression during sporozoite ontogeny and binding to human hepatocytes , 1995 .

[29]  T. Daly,et al.  Functional Characterization of a Redundant Plasmodium TRAP Family Invasin, TRAP-Like Protein, by Aldolase Binding and a Genetic Complementation Test , 2008, Eukaryotic Cell.

[30]  R. Ménard,et al.  Plasmodium sporozoite invasion into insect and mammalian cells is directed by the same dual binding system , 2002, The EMBO journal.

[31]  R. Hayward,et al.  The Plasmodium TRAP/MIC2 family member, TRAP‐Like Protein (TLP), is involved in tissue traversal by sporozoites , 2008, Cellular microbiology.

[32]  K. Sugamura,et al.  Molecular cloning of a glycosylphosphatidylinositol-anchored molecule CDw108. , 1999, Journal of immunology.

[33]  R. Medzhitov,et al.  Semaphorin 7A is a negative regulator of T cell responses. , 2006, Immunity.

[34]  Jacques J. Peschon,et al.  Semaphorin 7A promotes axon outgrowth through integrins and MAPKs , 2003, Nature.

[35]  L. Sibley,et al.  Aldolase forms a bridge between cell surface adhesins and the actin cytoskeleton in apicomplexan parasites. , 2003, Molecular cell.

[36]  K. Garcia,et al.  Structural Basis of Semaphorin-Plexin Recognition and Viral Mimicry from Sema7A and A39R Complexes with PlexinC1 , 2010, Cell.

[37]  R. Ménard,et al.  Conservation of a Gliding Motility and Cell Invasion Machinery in Apicomplexan Parasites , 1999, The Journal of cell biology.

[38]  Unified nomenclature for the semaphorins/collapsins. Semaphorin Nomenclature Committee. , 1999, Cell.

[39]  A. Crisanti,et al.  Identification of heparin as a ligand for the A-domain of Plasmodium falciparum thrombospondin-related adhesion protein. , 1999, Molecular and biochemical parasitology.

[40]  R. Sinden,et al.  CTRP is essential for mosquito infection by malaria ookinetes , 1999, The EMBO journal.

[41]  S. Thiberge,et al.  TREP, a novel protein necessary for gliding motility of the malaria sporozoite. , 2009, International journal for parasitology.

[42]  P. Permi,et al.  The layered fold of the TSR domain of P. falciparum TRAP contains a heparin binding site , 2006, Protein science : a publication of the Protein Society.

[43]  R. Coppel,et al.  No TRAP, no invasion. , 2009, Trends in parasitology.

[44]  G. Scott,et al.  Semaphorin 7a promotes spreading and dendricity in human melanocytes through beta1-integrins. , 2008, The Journal of investigative dermatology.

[45]  Andrea Crisanti,et al.  TRAP Is Necessary for Gliding Motility and Infectivity of Plasmodium Sporozoites , 1997, Cell.

[46]  A. Cowman,et al.  Plasmodium falciparum erythrocyte invasion through glycophorin C and selection for Gerbich negativity in human populations , 2003, Nature Medicine.

[47]  Amit Kumar Sharma,et al.  Role of Plasmodium falciparum thrombospondin-related anonymous protein in host-cell interactions , 2008, Malaria Journal.

[48]  Dominic P. Kwiatkowski,et al.  BASIGIN is a receptor essential for erythrocyte invasion by Plasmodium falciparum , 2011, Nature.

[49]  Dave Richard,et al.  A Conserved Molecular Motor Drives Cell Invasion and Gliding Motility across Malaria Life Cycle Stages and Other Apicomplexan Parasites* , 2006, Journal of Biological Chemistry.