Two genes encoding membrane antigens of Plasmodiumfalciparum were isolated by transient expression in mammalian cells and selection with human immune sera from African adults exposed to P. falciparum malaria. COS-7 cells were transfected with a plasmid expression library constructed from P. falciparum genomic DNA, and cells expressing reactive malaria antigens on their surface were enriched by adherence to antibody-coated dishes. One of the genes isolated is distinctive in that it does not contain repeat sequences typical of many malarial genes cloned by immunoscreening of bacterial expression libraries. The second gene apparently encodes a polymorphic version of the P. falciparum merozoite surface antigen Ag513, since the two sequences are identical in the 5' and 3' coding regions but diverge completely in the center. The COS-7 expression system provides an alternate means for cloning genes encoding malarial membrane antigens by using those antibodies in complex immune sera that bind membraneassociated, nondenatured molecules.Proteins expressed on the surface of parasites constitute potential target antigens for vaccine development. These molecules are accessible to the immune system and also may mediate functions vital for parasite survival such as cell attachment, host-cell invasion, or membrane transport. In the case of Plasmodium falciparum, surface membrane antigens of the asexual blood stages are of special interest because these stages are responsible for the high morbidity and mortality of P. falciparum malaria. The P. falciparum genes characterized to date have been cloned largely in bacterial expression systems, typically by using Agtll cDNA or genomic libraries in Escherichia coli and screening with human immune serum (1, 2). However, with these methods few genes for malarial surface antigens have been cloned, possibly due to poor expression of membrane proteins in E. coli and the fact that bacterial fusion proteins lack epitopes dependent on native secondary or higher order protein conformations. In this paper we show that genes encoding parasite membrane proteins can be isolated directly by expression in mammalian cells and selection for cells bearing heterologous surface antigens by using human immune serum from individuals exposed to the infection of interest. This method should be generally applicable to a number of parasitic diseases. Although the genes for a number of different human cell surface antigens have been cloned by transient expression in mammalian cells, these have been isolated exclusively from cDNA libraries, and immunological selection has depended on specific monoclonal antibodies (3-6). We demonstrate that genomic libraries can also be used in similar expression cloning and that polyspecific antibodies from human serum can be used for selection of transfected cells. MATERIALS AND METHODScDNA Expression Vector pJFE14. This simian virus 40 (SV40)-based COS cell expression vector is derived from pSSD2. The vector pSSD2 is similar to the original Okayama and Berg vector pcD...
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