A large body of evidence in rodent models and in human volunteers shows that protection against infection by malaria sporozoites can be obtained by vaccines containing the circumsporozoite (CS) protein, the major surface protein of sporozoites. Through its region II-plus motif, CS protein participates in the attachment step of the invasion of hepatocytes by the parasites. The investigator finds that another sporozoite membrane protein, the thrombospondin-related anonymous protein (TRAP), is additionally required for infectivity: TRAP (-) parasites do not exhibit the characteristic gliding motility of Apicomplexa, and do not infect the salivary glands of mosquitoes or the liver of the mammalian host. The TRAP gene encodes a typical membrane protein with a peptide anchor and a cytoplasmic domain. The N-terminal region of TRAP contains an approximately 220 residue module that is homologous to the A domains (or I domain of integrins) found in one or more copies in a large number of proteins involved in cell-cell, cell-matrix interactions. The high resolution crystal structure of A domains of integrins reveals the presence of a surface exposed metal ion dependent adhesion site (MIDAS) that participates in the recognition of the protein ligands. The long term objectives are to analyze the structural/functional properties of TRAP, and to use this information to improve the efficacy of CS vaccines.
The specific aims are 1) to obtain by gene targeting Plasmodium berghei lines with subtle mutations in TRAP of amino acids that are presumed to coordinate metal ion, and P. berghei lines with deletions in the cytoplasmic domain of TRAP. 2) to study the phenotype of these mutant parasites in the mosquito vector and mammalian host. 3) to express native and mutant forms of the A domain of P. berghei TRAP, study their ability to bind metal ions, and to inhibit sporozoite invasion of hepatocytes in vitro. 4) to develop synthetic vaccines that elicit specific antibodies against surface-exposed loops presumably located at the edges of the MIDAS of the A domain of P. yoelii TRAP.
